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ON 


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LIBRARY 


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DIRECTOK  AT  THE  CHEMICAL  LABORATORY  AT  THE  SOEBONNE 


mri/  TWENTY-EIGHT  ILLUSTRATIONS 


NEW  YORK : 
D.   APPLETON   AND  COMPANY, 
549  AND  551  Broadway. 
1876. 


CONTENTS. 


PAGE 

Introduction   i 

BOOK  I. 

FERMENTATION  DUE  TO  CELLULAR  ORGANISMS,  OR  DIRECT 
FERMENTATION. 

CHAPTER  I. 

Historical  9 

CHAPTER  II. 

Alcoholic  or  Spirituous  Fermentation     .      .  .18 

CHAPTER  III. 
AlcohqucJjerments  •      •  .34 

CHAPTER  IV. 

--N  Actual  Composition  of  Ferments  63 

CHAPTER  V. 

Functions  of  Yeast  72 

CHAPTER  VI. 
Action  of  various  Chemical  and  Physical  Agents 

on  Alcoholic  Fermentation   .      .      .      .  .159 

CHAPTER  VII. 
Can  nothing  but  Alcoholic  Yeast  excite  Alcoholic 

Fermentation   167 


vi 


CONTENTS. 


CHAPTER  VIII. 

PAGE 

Viscous  or  Mannitic  Fermentation  of  Sugar  .  .189 

CHAPTER  IX. 
Lactic  Fermentation   .193 

CHAPTER  X. 

Ammoniacal  Fermentation  203 

CHAPTER  XI. 
Butyric  Fermentation  and  Putrefaction  .      .      ,  209 

CHAPTER  XII. 
Fermentation  by  Oxidation  228 

CHAPTER  XIII. 
Application  of  the  Researches  and  Ideas  of  M. 
Pasteur  245 


BOOK  II. 

albuminoid  substances-soluble  or  indirect  ferments- 
origin  OF  ferments. 

CHAPTER  I. 

Albuminoid  Substances,  or  Proteids   •      •      •      .  253 
CHAPTER  II. 

Soluble  Ferments,  and  Indirect  Fermentation      .  269 

CHAPTER  III. 
On  the  Origin  of  Ferments  308 


LIST    OF  ILLUSTRATIONS. 


PAGE 

Fig.  I. — Saccharomyces    cerevisias  —  Sedimentary  yeast, 

X  400  diams.  45 

Fig.  2. — Saccharomyces  cerevisias — Sedimentary  yeast,  bud- 
ding, X  400  diams  47 

Fig.  3. — Saccharomyces  cerevisiae — Surface  yeast,  budding, 

X  400  diams.  47 

Fig.  4. — Saccharomyces  cerevisise — Surface  yeast,  at  rest, 

X  400  diams.  49 

Fig.  5. — Saccharomyces  cerevisiae — Sedimentary  yeast,  in  a 

growing  state,  X  400  diams  49 

Fig.  6. — Saccharomyces  cerevisiae — Formation  of  spores, 
X  750  diams.  a,  successive  phases  of 
the  production  of  spores  51 

Fig.  7. — Triads  of  spores,  germinating,  X  750  diams.    .       .  51 

Fig.  8. — Saccharomyces  elHpsoideus,  in  process  of  budding, 

X  600  diams.  55 

Fig.  9. — Saccharomyces  ellipsoideus,  development  of  spores, 

X  400  diams.  55 

Fig.  10. — Saccharomyces  elHpsoideus,  group  of  spores  in  the 

act  of  germinating,  X  400  diams  55 

Fig.  II. — Saccharomyces  Pastorianus,  X  400  diams.     •       .  56 

Fig.  12.— Saccharomyces  exiguus,  X  350  diams.    ...  56 

Fig.  13. — Saccharomyces  conglomeratus,  X  600  diams. .      .  56 


Vlii  LIST  OF  ILLUSTRATIONS. 

PAGE 


Fig.  14. — Saccharomyces    apiculatus   (Rees).  Carpozyma 

apic.  (Engel),  apiculated  ferment,  X  600  diams.  57 
Fig.  15. — Saccharomyces  Reesii,  ferment  of  red  wine,  X  350 

diams   59 

Fig,  16. — Saccharomyces  mycoderma,  X  350  diams.     •       ,  59 

Fig.  17. — Saccharomyces  mycoderma   59 

Fig.  18. — Mucor  racemosus,  ferment  in  mass        ...  60 
Fig.  19. — Apparatus  for  the  measurement  of  oxygen  dis- 
solved in  waters   121 

Fig.  20. — Viscous  Ferraents  of  Wine   190 

Fig.  21. — Lactic  Ferment      .   198 

Fig.  22. — Mycoderma  aceti   239 

Fig.  23. — Giret  and  Vinas'  apparatus  for  warming  wines  .  247 
Figs.  24  and  2 5. — Organic  corpuscules  of  dust,  mixed  with 

amorphous  particles   317 

Fig.  26. —  M.  Pasteur's  apparatus  for  the  introduction  of  cal- 
cined air  into  flasks  containing  organic  infusions  319 


Fig.  27. — M.  Pasteur's  apparatus  for  studying  the  resistance 
of  germs  and  spores  to  temperatures  more  or  less 
elevated  323 

Fig.  28. — M.  Pasteur's  flask  to  deprive  the  air  of  its  germs  .  323 


NOTE  BY  TRANSLATOR. 

The  French  words  "  invertir,"  "inverti,"  "  intervertir,"  "inversive,"  &c., 
p.  28,  &c.,  may  be  rendered  in  English  by  the  recognized  terms,  "inverted," 
*'  inversive,"  &c.  Yet  since  these  terms  are  not  understood  by  all,  and  present 
some  difficulty,  it  has  been  thought  better  to  adopt  in  the  text  the  less  technical 
and  more  suggestive  rendering  "altered,"  "  alterative," '  &c. ;  by  altered  sugar 
being  meant  cane-sugar  which  has  taken  up  a  molecule  of  water  and  split  up 
into  a  mixture  of  glucose  and  levulose. 


ON  FERMENTATION. 


INTRODUCTION. 

F'ERMENTATION  is  only  a  particular  instance,  selected 
from  among  the  chemical  phenomena  of  which  living 
organisms  are  the  field  ;  it,  like  all  biological  reactions, 
comes  before  us  as  a  manifestation  of  the  special  force 
residing  in  these  organisms,  or  rather  in  their  cellular 
elements. 

If  we  leave  the  nature  of  the  fermenting  body,  and 
the  products  derived  from  it,  in  the  background, 
there  is  nothing  to  distinguish  fermentation  from  the 
other  chemical  transformations  which  take  place  in  the 
animal  or  vegetable  economy. 

The  reason  why  the  production  of  alcohol  and  carbon 
dioxide  at  the  expense  of  sugar,  the  conversion  of 
glucose  into  lactic  and  butyric  acids,  and  other  pheno- 
mena of  the  same  order  have  been  classed  by  them- 
selves, is  that  the  real  cause  of  these  curious  transfor- 
mations was  long  misunderstood.  It  had  not  been 
observed  that  they  had  as  their  origin  the  presence  of 
living  organisms,  or,  at  least,  principles  which  are 
directly  derived  therefrom. 


ON  FERMENTATION. 


There  is,  therefore,  no  longer  any  necessity,  in  the 
present  state  of  science,  for  grouping  together  under  a 
special  name  these  various  reactions  ;  it  is  more  con- 
venient, on  the  contrary,  to  class  them  among  the 
general  mass  of  chemical  phenomena  of  the  living 
organism. 

We  must,  consequently,  do  one  of  two  things  ;  either 
cease  to  use  the  term  fermentation,  as  a  general  ex- 
pression applying  to  a  certain  order  of  phenomena, 
or  we  must  designate  by  it  all  those  changes  which, 
by  the  special  conditions  under  which  they  are  pro- 
duced, are  evidently  due  to  the  intervention  of  a 
force  differing  from  those  which  we  handle  in  our  labo- 
ratories. 

It  is  true  that  the  organisms  which  give  rise  to  what 
have  been  hitherto  called  fermentation  are  simple 
elementary  organisms  reduced  to  a  single  cell  ;  but  a 
plant  or  an  animal  of  a  high  order  is  only  the  union, 
under  special  laws,  of  different  kinds  of  cells,  each  of 
which  acts  in  a  certain  determinable  manner.  When, 
as  M.  Pasteur  has  remarked,  we  sow  at  the  same  time, 
in  the  same  saccharine  medium,  alcoholic,  lactic,  and 
butyric  ferment,  we  see  three  distinct  reactions  take 
place,  one  of  which  splits  up  the  sugar  into  alcohol  and 
carbon  dioxide,  the  second  converts  it  into  lactic  acid, 
and  the  third  into  butyric  acid. 

The  more  simple  an  organism  is,  the  fewer  special 
kinds  of  cells  it  contains,  the  simpler  are  the  chemical 
reactions  which  take  place  in  it,  and  the  more  ear>ily  are 
they  separated  from  each  other,  and  isolated  by  experi- 
ment. On  the  contrary,  in  proportion  as  the  histological 
constitution  is  varied  and  heterogeneous,  we  see  a 


INTRODUCTION. 


3 


greater  number  of  distinct  compounds,  as  the  products 
of  the  many  chemical  changes  which  take  place  in  the 
different  tissues. 

As  a  consequence  of  what  we  have  just  said,  our  plan 
would  be  considerably  enlarged,  and  the  history  of 
fermentation  would  become  that  of  the  chemical  phe- 
nomena of  life. 

We  will  not,  however,  give  such  a  wide  scope  to  this 
w^ork,  but  will  confine  ourselves  to  the  examination  of 
the  phenomena  which  have  been  hitherto  designated  by 
the  name  of  fermentation.  Under  these  restrictions,  the 
history  of  fermentation  may  be  considered  as  an  intro- 
duction to  biological  chemistry. 

In  fact,  it  is  easily  seen,  from  the  preceding  consi- 
derations, that  the  thorough  study  of  ferments,  properly 
so  called,  or  rather  of  elementary  organisms,  and  of 
their  mode  of  existence,  ought  to  precede  that  of  the 
more  complete  beings.  We  more  easily  understand  the 
properties  of  granite,  and  the  influence  exerted  upon  it 
by  water  and  atmospheric  agents,  when  we  have  learned 
that  it  is  formed  of  crystals  of  quartz,  felspar,  and  mica  in 
juxtaposition,  and  have  studied  the  chemical  characters 
of  each  of  these  compounds.  In  the  same  manner,  the 
study  of  the  chemical  manifestations  of  the  vital  force 
in  cellular  organisms  is  destined  to  throw  a  bright  light 
on  the  more  complex  functions  of  the  higher  plants  and 
animals.  This  has  been  recognized  by  M.  Pasteur,  and 
by  all  those  who  have  subsequently  entered  on  the 
physiological  study  of  fermentation,  and  of  the  develop- 
ment of  cellular  organisms. 

A  living  cell  of  beer-yeast  possesses  the  property  of 
resolving  into  alcohol,  glycerin,  carbon  dioxide,  and 


4 


ON  FERMENTATION. 


succinic  acid  the  altered  sugar  which  penetrates  by 
endosmose  through  its  membranous  envelope. 

If  we  substitute  for  the  cell  of  beer-yeast  a  cell  of 
lactic  ferment,  we  still  see  the  sugar  disappear,  but  the 
products  into  which  the  ponderable  elements  of  the 
glucose  are  resolved  are  different ;  instead  of  alcohol 
and  carbon  dioxide,  we  have  lactic  acid.  The  modus 
faciendi  of  the  vital  force  of  this  cell  is  evidently  not 
the  same  as  that  of  the  former  one  ;  but  we  cannot, 
therefore,  affirm  that  there  are  as  many  vital  chemical 
forces  as  there  are  reactions. 

When  a  pencil  of  solar  light  passes  through  a  prism, 
the  constituent  parts  of  this  pencil  are  isolated  on 
account  of  their  unequal  refrangibility.  The  least 
refrangible  rays  are  revealed  to  us  by  the  effects  of 
heat  (the  dilatation  and  change  of  condition  of  bodies) ; 
next  come  the  luminous  rays,  which  excite  on  the  retina 
the  impressions  of  colour  forming  the  spectrum  ;  and 
then,  beyond  the  violet,  is  a  series  of  invisible  rays, 
which  are  revealed  only  by  their  decomposing  action  on 
certain  combinations  (salts  of  silver,  &c.).  But  we 
know  now  that  all  these  calorific,  luminous,  and  chemi- 
cal rays,  some  of  which  give  heat  without  light,  and 
others  light  without  heat,  or  excite  chemical  reactions, 
differ  only  in  the  rapidity  of  the  vibratory  movements  of 
the  ether,  and  are  essentially  distinguishable  from  each 
other  only  by  their  wave-length.  It  is  possible  that  an 
analogous  bond  may  unite  the  vital  chemical  forces  of 
the  different  elementary  organisms.  Sand,  sprinkled 
uniformly  on  the  surface  of  a  vibrating  plate,  collects 
in  nodal  lines  of  different  forms,  according  to  the  sharp- 
ness of  the  note  which  we  draw  from  this  plate  by 


INTRODUCTION. 


5 


means  of  a  bow ;  in  the  same  manner,  chemical  com- 
pounds may  perhaps  be  resolved  into  more  simple 
combinations,  varying  in  kind  according  to  the  vibratory 
rhythm  which  starts  them. 

The  transformation  of  sugar  into  alcohol  and  carbon 
dioxide,  and  the  conversion  of  the  same  body  into 
lactic  acid  are  chemical  phenomena  which  we  cannot  yet 
reproduce  by  the  intervention  of  heat  alone,  nor  by  the 
additional  agency  of  light  or  of  electricity.  The  force 
capable  of  attacking,  in  a  certain  determinate  direction, 
the  complex  edifice  which  we  call  sugar,  an  edifice 
composed  of  atoms  of  carbon,  hydrogen,  and  oxygen, 
grouped  according  to  a  determinate  law — this  force, 
which  is  manifested  only  in  the  living  cell  of  the  fer- 
ment, is  a  force  as  material  as  all  those  which  we  are 
accustomed  to  utilize. 

Its  principal  peculiarity  is,  that  it  is  only  found  in  the 
living  organisms,  to  which  it  gives  their  peculiar  charac- 
ter. We  ought  not  to  allow  ourselves  to  be  stopped  by 
this  rampart,  over  which  no  one  has  hitherto  been  able 
to  pass  ;  we  ought  not  to  say  to  the  chemist,  You 
shall  go  no  farther,  for  beyond  this  is  the  domain  of 
life,  where  you  have  no  control." 

The  history  of  science  shows  us  the  weakness  of  these 
so-called  impassable  barriers. 

Gerhardt,  when  he  published  his  excellent  treatise 
on  organic  chemistry,  thought  himself  justified  in  say- 
ing, "  It  is  vital  force  alone  which  acts  synthetically 
and  reconstructs  the  edifice  demolished  by  chemical 
forces." 

M.  Berthelot,  some  years  afterwards,  in  a  brilliant 
series  of  discoveries,  made  the  first  successful  attempt  to 


6 


ON  FERMENTATION. 


perform  organic  syntheses,  and  determined  the  principal 
conditions  under  which  they  can  be  effected. 

In  a  remarkable  lecture  on  molecular  dissymmetry 
(Lemons  de  la  Societe  Chimique  de  Paris,  i860), 
M.  Pasteur  had  established  an  important  distinction 
between  artificial  organic  products  and  the  compounds 
formed  under  the  influence  of  living  organisms. 

"  The  artificial  products  of  the  laboratory  have  coinci- 
dent images  (sont  a  image  superposable).  On  the  con- 
trary, most  of  the  natural  organic  products — I  might  say 
all^  if  I  had  only  to  allude  to  those  which  play  an  impor- 
tant part  in  the  phenomena  of  vegetable  and  animal  life — 
all  the  products  essential  to  life  are  unsymmetrical,  and 
unsymmetrical  in  such  a  way  that  their  images  cannot 
be  made  to  coincide  with  them."  And  afterwards  he 
says,  We  have  not  yet  been  able  to  realize  the  produc- 
tion of  an  unsymmetrical  body,  by  the  aid  of  compounds 
which  are  not  so  themselves." 

Nearly  at  the  same  time  that  these  words  were 
uttered  before  the  Chemical  Society  of  Paris,  two 
English  chemists,  Perkin  and  Duppa,  succeeded  in 
transforming  succinic  acid  into  tartaric  acid.  M. 
Pasteur  himself  acknowledged  that  the  artificial  pro- 
duct of  Perkin  was  a  mixture  of  paratartaric  acid  and 
of  inactive  tartaric  acid.  But  paratartaric  acid  easily 
splits  up,  as  Pasteur's  elegant  experiments  have  shown, 
into  dextro-tartaric  and  la^vo-tartaric  acid,  and  M. 
Jungflcisch  has  shown  that  inactive  tartaric  acid,  heated 
with  water  at  175°,  is  partially  converted  into  para- 
tartaric acid. 

The  succinic  acid  employed  by  the  l^nglish  chemists 
was  formed  by  the  oxidation  of  yellow  amber.  This 


INTRODUCTION. 


was  not  a  synthetical  product ;  it  might  be  thought 
that,  though  it  was  inactive,  it  resulted,  like  racemic 
acid,  from  the  union  of  two  active  but  opposed  mole- 
cules. Jungfleisch  has  removed  this  last  doubt.  He 
prepared  synthetic  succinic  acid  by  a  well-known 
method,  by  means  of  cyanide  of  ethylene  and  potassium. 
This  acid  furnished  paratartaric  acid,  like  that  produced 
from  amber. 

Thus  fell  the  barrier  placed  by  M.  Pasteur  between 
natural  and  artificial  products.  This  example  shows  us 
how  cautious  we  ought  to  be  in  making  distinctions 
which  we  seem  justified  in  establishing  between  the 
chemical  reactions  of  the  living  organism  and  those  of 
the  laboratory.  Because  a  chemical  phenomenon  may 
hitherto  have  been  produced  only  under  the  influence  of 
life,  it  does  not  follow  that  it  will  never  be  effected 
otherwise. 

No  one  can  any  longer  admit  that  vital  force  has 
power  over  matter,  to  change,  counterbalance,  or  annul 
the  natural  play  of  chemical  affinities.  That  which  we 
have  agreed  to  call  chemical  affinity  is  not  an  absolute 
force  ;  this  affinity  is  modified  in  numberless  ways, 
according  as  the  circumstances  by  which  bodies  are 
surrounded,  vary.  Thus,  the  apparent  differences 
between  the  reactions  of  the  laboratory  and  those  of 
the  organism  ought  to  be  sought  for,  more  particularly 
among  the  special  conditions,  which  the  latter  alone  has 
been  able  hitherto  to  bring  together. 

In  other  words,  there  is  really  no  chemical  vital  force. 
If  living  cells  produce  reactions  which  seem  peculiar 
to  themselves,  it  is  because  they  realize  conditions  of 
molecular  mechanism  which  we  have  not  hitherto  sue- 


8 


ON  FERMENTATION. 


ceeded  in  tracing,  but  which  we  shall,  without  doubty  be 
able  to  discover  at  some  future  time.  Science  can 
gain  nothing  by  being  limited  in  the  possibility  of  the 
aims  which  she  proposes  to  herself,  or  the  end  which 
she  seeks. 

If,  in  this  work,  we  still  employ  the  expression,  the 
vital  chemical  force  of  an  elementary  organism,"  it  will 
be  clearly  understood  that  we  intend  these  words  to 
signify  the  realization  of  the  conditions  of  molecular 
mechanism  necessary  in  order  to  set  up  a  certain  reaction. 

We  will  not  delay  any  longer  over  these  general 
considerations,  which  are,  after  all,  nothing  but  hypo- 
theses, naturally  suggesting  themselves  to  the  mind  of 
him  who  seeks  to  explain  the  causes  which  produce 
certain  observed  effects,  but  on  which  it  is  not  neces- 
sary to  dwell  at  the  present  stage  of  our  inquiry ;  we 
will  therefore  proceed  at  once  to  the  examination  of 
facts. 

The  study  of  fermentation  may  be  divided  into  two 
parts,  according  to  the  nature  of  the  ferment.  The 
first  will  comprise  the  fermentation  due  to  the  inter- 
vention of  an  organized  ferment,  having  a  determinate 
form ;  the  second  will  be  reserved  for  fermentation 
produced  by  soluble  products,  elaborated  by  living 
organisms. 


BOOK  I. 

FERMENTATION  DUE  TO  CELLULAR  ORGANISMS.  OR 
DIRECT  FERMENTATION. 


CHAPTER  L 

HISTORICAL. 

The  word  lermentation  is  derived  ivom.  fervere^  to  boil; 
it  evidently  owes  its  origin  to  the  reaction  presented  by 
saccharine  liquids,  when  they  are  left  to  themselves  or 
placed  in  contact  with  ferments.  We  observe,  in  fact, 
in  this  case,  a  more  or  less  abundant  disengagement  of 
gas,  which  causes  the  liquid  to  effervesce  or  boil.  The 
sugar  disappears,  and  the  product  becomes  spirituous. 
The  expression,  fermentation,  was  subsequently  applied 
to  other  phenomena,  in  which  an  organic  body,  when 
dissolved,  is  modified,  changed,  and  transformed,  under 
the  influence  of  a  cause  which  remained  for  a  long  time 
unknown  and  badly  defined.  Thus  the  acidification  of 
wine  was  called  fermentation,  although  in  this  case 
there  was  no  effervescence.  The  analogy  of  the  deter- 
mining cause  was  considered,  rather  than  the  appear- 
ance of  the  phenomenon. 

Alcoholic  fermentation  was  the  first  known,  and  was 


lO  ON  FERMENTATION. 

also  more  studied  than  the  other  reactions  of  this  class. 
Osiris  among  the  Egyptians,  Bacchus  among  the 
Greeks,  Noah,  according  to  the  IsraeHtish  tradition, 
taught  men  the  art  of  cultivating  the  vine,  and  making 
wine.  Moses,  in  his  writings,  draws  a  distinction  be- 
tween unleavened  and  leavened  bread,  and  relates  that 
the  Israelites  were  in  such  haste,  during  their  flight  from 
Egypt,  that  they  had  no  time  to  put  leaven  into  their 
dough.  The  ancients  used  as  leaven  for  their  bread 
either  dough  that  had  been  kept  till  it  was  sour,  or  beer- 
yeast. 

"Galliae  et  Hispania^  frumento  in  potum  resoluto, 
spuma  ita  concreta  pro  fermento  utuntur,  qua  de  causa 
levior  illis,  quam  ceteris,  panis  est,"  says  Pliny,  who 
also  adds,  that  in  the  fermentation  of  bread  acidity  is 
the  most  active  principle.  From  the  earliest  times  cer- 
tain fermented  liquids  were  known,  both  in  Egypt  and 
Germany,  prepared  from  natural  saccharine  juices  which 
had  been  allowed  to  ferment ;  such  as  beer,  hydromel, 
palm-wine,  and  cider. 

We  find,  in  short,  from  all  ancient  documents,  that 
alcoholic  fermentation  was  empirically  known  in  its 
principal  effects,  and  utilized  at  a  period  far  earlier  than 
that  which  has  left  written  traces  of  its  history. 

Among  the  writings  of  the  alchemists  from  the 
thirteenth  to  the  fifteenth  century,  we  very  frequently 
find  the  expressions  "  fermentation  and  ferments  " 
(fcrmcntatos  et  fcrmentum),  without  our  being  able  to 
ascertain  clearly  what  precise  ideas  they  attached  to 
them.  They  knew  no  distinction  between  mineral  and 
organic  substances ;  the  phenomena  connected  with 
the  changes  in  organic  products  were  assimilated  and 


HISTORICAL. 


II 


confounded  with  the  transformations  of  mineral  com- 
pounds, and  with  the  solution  of  salts  and  metals.  The 
term  "  ferment "  was  often  applied  even  to  the  philo- 
sopher's stone. 

"  Apud  philosophos  fermentum  dupliciteo  videtur  dici ; 
uno  modo  ipse  lapis  philosophorum  et  suis  dementis 
compositus,  et  completus  in  comparatione  ad  metalla ; 
alio  modo  illud,  quod  est  perficiens  lapidem  et  ipsum 
complens. 

"De  primo  modo  dicimus,  quod  sicut  fermentum 
pastse  vincit  pastam,  et  ad  se  convertit  semper,  sic  et 
lapis  convertit  ad  se  metalla  reliqua.  Et  sicut  una  pars 
fermenti  pastae  habet  convertere  partes  pastse  et  non 
converti,  sic  et  hie  lapis  habet  convertere  plurimas 
partes  metallorum  ad  se,  et  non  converti.'* — Petrus 
Bonus  of  Ferraruy  1 330-1 340. 

We  see  that  the  writer  is  especially  struck  with  this 
fact,  that  a  very  small  quantity  of  leaven  transforms 
into  fresh  leaven  an  almost  indefinite  quantity  of  paste. 
This  property  of  transmitting  a  force  to  a  large  mass 
without  being  itself  weakened  by  the  process,  was  pre- 
cisely that  which  ought  to  characterize  the  philosopher's 
stone  which  was  so  much  sought  after. 
.  Basil  Valentine,  in  his  Triumphal  Car  of  Antimony," 
admits  that  yeast,  employed  in  the  preparation  of  beer, 
communicates  to  the  liquor  an  internal  inflammation, 
and  determines  thereby  a  purification,  and  a  separation 
of  the  clear  parts  from  those  which  are  troubled. 

Alcohol,  the  presence  of  which  in  the  fermented 
liquid  was  known  to  him,  was  considered  by  him  to 
exist  previously  in  the  decoction  of  germinated  barley  ; 
but  that  it  did  not  become  active  and  susceptible  of 


12 


ON  FERMENTATION. 


being  separated  by  distillation  until  it  had  been  cleared 
from  the  impurities  which  accompany  it,  and  mask  its 
special  properties. 

Libavius  (Alchymia,  1595)  believed  that,  "Fermen- 
tatio  est  rei  in  substantia,  per  admistionem  fermente 
quod  virtute  per  spiritum  distributo  totam  penetrat 
massam  et  in  suam  naturam  immutat,  exaltatis."  The 
ferment  must  be  of  a  similar  nature  to  the  matter  which 
enters  into  fermentation,  and  the  latter  must  be  either 
liquid,  or  in  a  state  of  minute  division  ;  the  principal 
agent  resides  in  the  heat  of  the  ferment. 

Like  the  chemists  of  a  later  age,  Libavius  compares 
fermentation  to  putrefaction,  and  considers  them  as  dif- 
ferent effects  of  the  same  cause.  He  protests,  on  the 
contrary,  against  the  confused  ideas  which  had  been 
entertained  concerning  digestion  and  fermentation. 
Digestion  is,  according  to  him,  "motus  ad  mistionem, 
non  ad  perfectionem,"  as  fermentation  is. 

The  iatro-chemical  school  attributed  to  fermentation 
a  preponderating  power,  and  even  confounded  under  this 
term  a  great  number  of  chemical  reactions. 

Thus  Van  Helmont  expresses  himself  as  follows  in 
his  "  Ortus  Medicinae  "  (648) :  "  Docebo  omnem  trans- 
mutationem  formalem  prsesupponere  fermentum  cor- 
ruptivum." 

The  formation  of  intestinal  gasses,  the  production  of 
blood  and  animal  fluids,  spontaneous  generation,  the 
effervescence  of  chalk  under  the  influence  of  acids, 
arc  phenomena  with  which  fermentation  has  to  do. 

We  may,  however,  say  in  passing  that  Van  Helmont 
had  the  merit  of  clearly  distinguishing  the  production 
of  a  special  gas  (gas  vinorum)  during  alcoholic  fermen- 


HISTORICAL. 


13 


tation.  He  says  that  this  gas  vinorum  is  different  from 
spirit  of  wine,  as  he  was  able  to  prove  by  experiments. 
It  is  impossible  to  discover  from  his  writings  whether 
or  no  he  recognized  the  identity  of  the  gas  vinorum  and 
the  gas  carbonum  produced  during  the  combustion  of 
charcoal. 

In  1664,  Wren  pointed  out  that  the  gas  produced  by 
alcoholic  fermentation  can  be  absorbed  by  water,  like 
that  which  is  disengaged  by  the  action  of  an  acid  on 
salt  of  tartar. 

Silvius  de  la  Boe  (1659)  no  longer  regarded  the  effer- 
vescence of  the  alkaline  carbonates  under  the  influence 
of  acids  as  a  phenomenon  of  the  same  class  as  fermen- 
tation. 

He  supposed  that  in  the  former  case  there  was  com- 
bination, in  the  latter  decomposition. 

Lemery  (Cours  de  Chimie,  1675)  is  not  so  explicit 
when  he  says:  "The  fermentation  which  occurs  in  paste, 
wort,  and  all  other  similar  things,  is  different  from  that 
of  which  we  have  just  spoken  (effervescence),  since  it  is 
slower  ;  it  is  excited  by  the  natural  acid  salt  of  these 
substances,  which,  becoming  disengaged,  and  having  its 
energy  increased  by  its  motion,  rarefies  and  raises  the 
gross  and  oily  part  which  opposes  its  passage,  and  thus 
we  see  the  matter  rise. 

The  reason  why  the  acid  does  not  cause  sulphurous 
substances  to  ferment  with  as  much  noise  and  readiness 
as  alkalis,  is  that  oils  are  composed  of  pliant  parts  which 
yield  to  the  points  of  the  acid,  as  a  piece  of  wool  or  cotton 
would  yield  to  needles  pressed  into  it.  Thus  it  seems 
to  me  that  we  must  admit  of  two  sorts  of  fermentations  ; 
one  of  acids  with  alkalis,  which  would  be  called  effer- 


14 


ON  FERMENTATION. 


vescence  ;  and  the  other  would  be,  when  the  acid  rarefies 
by  degrees  a  solid  matter  like  paste,  or  clear  and 
sulphurous  like  wort,  cider,  or  other  juices  of  plants ; 
we  should  call  the  latter  sort  fermentation." 

Lemery  says  again,  when  speaking  of  alcoholic  fermen- 
tation :  "  In  order  to  understand  this  effect,  we  must 
know  that  wort  contains  much  essential  salt ;  this  salt, 
being  volatile,  makes  an  effort,  during  fermentation,  to 
detach  itself  from  the  oily  particles  by  which  it  is,  as  it 
were,  bound  ;  it  penetrates  them,  divides  and  separates 
them,  until  by  its  subtile  and  piercing  points,  it  has  rare- 
fied them  into  spirit;  this  effort  causes  the  ebullition 
which  takes  place  in  wine,  and,  at  the  same  time,  its 
purification ;  for  it  separates  and  removes  the  grosser 
parts  in  the  form  of  froth,  a  portion  of  which  attaches 
itself  to  the  sides  of  the  vessel  and  grows  hard  ;  the 
other  falls  to  the  bottom,  and  is  called  tartar  and  lees. 
The  inflammable  spirit  of  wine  is  therefore  nothing  but 
an  oil  exalted,  that  is  purified,  by  salt.'* 

We  find  in  the  researches  and  writings  of  Becker 
(1682),  a  very  marked  progress  in  the  study  of  the  pro- 
ducts of  fermentation.  He  was  the  first  to  bring  for- 
ward the  important  fact  that  saccharine  liquids  alone 
are  capable  of  entering  into  spirituous  fermentation. 
He  considers  that  alcohol  does  not  pre-exist  in  the 
wort,  but  is  formed  during  the  process  of  fermentation  ; 
the  intervention  of  air  is  necessary  to  set  this  action 
going,  which  he  considers  analogous  to  combustion. 
Becker  brings  together,  under  the  name  of  fermentation, 
the  production  of  gas  in  the  stomach  of  sick  animals 
(insumcfactio),  spirituous  fermentation  (proprie  fermen- 
tatio),  and  acetification  (acctificatio  scu  acesccntia). 


J 


HISTORICAL. 


15 


We  owe  to  Willis  (1659),  and  Stahl,  the  celebrated 
originator  of  the  theory  of  phlogiston  (1697),  the  first 
philosophical  conception  of  the  peculiar  nature  of  fer- 
mentation, or  rather  of  fermentations.  According  to 
their  views  a  ferment  is  a  body  endued  with  a  motion 
peculiar  to  itself,  and  it  transmits  this  motion  to  the 
fermentable  matter.  Thus  Willis  says  in  his  disserta- 
tion   De  Fermentatione": — 

"Fermentatio  est  motus  intestinus  cujusvis  corporis, 
cum  tendentia  ad  perfectionem  ejusdem  corporis  vel 
propter  mutationem  in  aliud.  Plures  sunt  modi  quibus 
fermentatio  promooctur.  Primus  et  principius  erit 
fermenti  cujusdam  corpori  fermentando  adjectio ;  cujus 
particulae  cum  prius  sint  in  vigore  et  motu  positae,  alias 
in  massa  fermentanda  otiosas  et  torpidas  exsuscitant,  et 
in  motum  vindicant." 

Stahl  considered  alcoholic  fermentation  as  a  pheno- 
menon of  the  same  class  as  putrefaction,  and  as  only 
a  particular  case  of  it.  As  there  was  at  that  time  no 
definite  idea  of  the  elementary  composition  of  ferment- 
able substances,  and  of  the  products  of  their  fermenta- 
tion, there  evidently  could  not  be  established  any  correct 
relation  between  these  bor'^'es,  and  any  hypothesis  could 
be  safely  brought  forward.  Thus  Stahl  considers  that 
fermentable  matter  (sugar,  flour,  milk)  is  composed  of 
particles  formed  by  the  unstable  union  of  salt,  oil,  and 
earth ;  under  the  influence  of  the  internal  motion  excited 
by  the  ferment,  the  heterogeneous  particles  are  separ- 
ated from  each  other,  and  then  recombined  so  as  to 
form  more  stable  compounds  including  the  same  prin- 
ciples, but  in  other  proportions. 

From  Stahl  to  Lavoisier  we  find  no  names  of  great 


i6 


ON  FERMENTATION. 


note,  and  no  interesting  discoveries  with  respect  to 
fermentation. 

When  chemistry  underwent  its  great  transformation 
at  the  end  of  the  last  century,  under  the  powerful  influ- 
ence of  the  genius  of  Lavoisier,  fermentation  necessa- 
rily attracted  anew  the  attention  of  experimentalists. 
Lavoisier  himself  studied  it  (Elemens  de  Chimie,  vol.  i. 
p.  139,  second  edition),  and  as  was  the  case  with  all 
subjects  which  he  handled,  he  threw  a  ray  of  light 
upon  the  darkness.  Proceeding  in  his  usual  manner, 
balance  in  hand,  by  weight  and  measure,  and  applying 
to  the  solution  of  the  problem  the  new  methods  of 
organic  analysis  which  he  had  invented,  he  endeavoured 
to  ascertain  the  bond  or  relation  which  exists  between 
the  fermented  matter,  the  sugar,  and  the  products  of 
fermentation,  alcohol  and  carbon  dioxide. 

From  this  moment  we  quit  the  domain  of  the  history 
of  the  science,  and  enter  into  that  of  the  real  and  well- 
observed  facts  which  will  be  treated  of  in  the  following 
chapters. 

We  may  say,  in  recapitulation,  that,  before  the  labours 
of  Lavoisier  and  his  followers,  the  fermentable  products 
and  the  principal  terms  of  their  transformations  (carbon 
dioxide  gas,  alcohol,  acetic  acid,  &c.)  were  known  qicali- 
tativcly.  The  distinction  between  the  acid,  or  acetic 
fermentation,  and  the  alcoholic  fermentation  was  known ; 
there  was  an  idea  of  the  analogy  which  exists  between 
putrefaction  and  alcoholic  fermentation ;  and  an  ex- 
planation of  the  manner  in  which  a  ferment  acts  had 
been  sought. 

The  latter  was  known  only  as  a  kind  of  foam,  deposit, 
or  paste,  in  which  resided  an  occult  and  special  force, 


HISTORICAL. 


17 


capable  of  determining  chemical  phenomena.  We  may 
add  that  these  phenomena  were  considered  as  distinct, 
both  in  their  action  and  exciting  cause,  from  the 
ordinary  reactions  of  chemistry.  This  was,  as  one  may 
see,  but  a  slight  result  of  the  many  volumes  that  had 
been  written  on  this  subject. 

Spirituous  or  alcoholic  fermentation  being,  in  every 
respect,  the  part  of  this  subject  which  has  been  the  most 
thoroughly  studied,  we  will  commence  our  monograph 
by  its  examination. 

2 


ON  FERMENTATTON. 


CHAPTER  II. 

ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION. 

Pasteur,  in  his  exgellent  memoir  (Ann.  de  Chimie  et 
Physique,  3rd  series,  vol.  Iviii.  p.  323),  calls  by  the  name 
of  alchoholic  fermentation  that  which  sugar  undergoes 
under  the  influence  of  the  ferment  which  bears  the 
name  of  yeast  or  barm. 

We  can  only  adopt  this  definition  as  applying,  with- 
out any  possibility  of  uncertainty,  to  a  phenomenon 
very  limited  in  its  cause  and  its  effects ;  but  we  shall 
have  to  inquire,  in  a  later  portion  of  this  work,  whether 
alcohol  cannot  be  produced  at  the  expense  of  sugar 
under  other  influences  than  those  of  the  product  known 
as  becr-yeast. 

As  we  have  before  said,  the  splitting  up  of  a  molecule 
of  sugar  into  many  more  simple  products,  among  which 
we  find  alcohol  and  carbon  dioxide,  is  the  consequence 
of  a  special  mechanical  action,  exercised  on  the  ultimate 
particles  of  the  compound  matter.  Whatever  may  be 
the  source,  whether  living  organism  or  dead  matter, 
which  realizes  the  conditions  necessary  for  this  rupture 
of  equilibrium,  the  phenomenon  will  be  essentially  the 
same.  In  a  general  and  philosophical  point  of  view, 
there  is  no  more  reason  why  we  should  separate  alco- 
holic fermentation  excited  by  yeast  from  that  which 
is   due  to  any   other  agent,   than    why  we  should 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.      1 9 


distinguish  cane  sugar  from  that  produced  from  beet- 
root. 

While  we  restrict,  with  Pasteur,  the  expression  "  alco- 
holic fermentation,"  and  do  not  include  in  it  all  the 
phenomena  of  decomposition,  in  which  alcohol  is  pro- 
duced, we  have  to  consider  the  body  which  ferments, 
the  sugar,  or  rather  the  sugars,  the  products  of  fermen- 
tation, among  which  alcohol  takes  its  place  in  the  first 
rank,  and  then  the  determining  cause  of  the  fermenta- 
tion of  sugar,  beer-yeast. 


Products  of  the  Reaction. 

Let  us  first  consider  alcoholic  fermentation  as  an 
ordinary  chemical  reaction  ;  in  other  words,  let  us  study 
it  by  means  of  the  body  which  is  decomposed,  and  the 
products  which  are  derived  from  it;  we  will  then 
consider  the  cause  of  the  decomposition,  and  the  pro- 
perties of  this  ferment,  as  well  as  those  of  analogous 
products. 

We  said  before  that  Becker  was  the  first  to  recognize 
the  necessity  of  the  presence  of  sugar  in  wines  which 
undergo  spirituous  fermentation,  but  that  to  Lavoisier 
belongs  the  honour  of  having  studied  and  demonstrated 
the  relations  of  their  composition  which  connect  sugar 
with  its  derivatives. 

Setting  out  with  this  principle,  that  nothing  is  created 
either  in  the  operations  of  art,  or  in  those  of  nature  ; 
that  in  every  operation  there  is  an  equal  quantity  of 
matter  both  before  and  after  the  operation  ;  that  the 


20 


ON  FERMENTATION. 


quality  and  quantity  of  the  elements  are  the  same,  and 
that  there  are  only  changes  and  modifications,  this 
illustrious  chemist  established  by  analysis  the  centesi- 
mal proportions  of  carbon,  hydrogen,  and  oxygen 
contained  in  sugar,  operating  in  the  same  manner  on 
the  alcohol,  the  carbon  dioxide,  and  acetic  acid  recog- 
nized by  him  as  the  products  of  the  decomposition  of 
sugar ;  then,  estimating  by  analysis  the  respective 
quantities  of  these  three  bodies  which  are  formed  at 
the  expense  of  a  known  weight  of  sugar,  he  ascertained 
the  result  of  the  reaction,  and  arrived  at  the  following 
conclusions : — 

"  The  effects  of  vinous  fermentation  are  reduced  to 
separating  into  two  portions  the  sugar,  which  is  an 
oxide,  and  oxidizing  one  at  the  expense  of  the  other, 
so  as  to  form  from  it  carbon  dioxide,  in  reducing 
the  other  in  favour  of  the  former,  to  form  from  it  a 
combustible  substance,  alcohol  ;  so  that  if  it  were  possi- 
ble to  recombine  these  two  substances,  alcohol  and 
carbon  dioxide,  we  should  reform  the  sugar."  He  had 
really  made  a  great  advance  on  the  conceptions  of 
Stahl,  founded  on  a  mixture  of  salt,  oil,  and  earth. 

The  researches  of  Lavoisier  may  be  summed  up  by 
the  following  equation  : — 

95*9  parts  of  crystallized  cane  sugar  contain  26*8  of 
carbon,  77  of  hydrogen,  and  61*4  of  oxygen. 

These  are  decomposed,  and  form  577  parts  of 
alcohol,  containing  167  carbon,  96  hydrogen,  and 
31*4  oxygen  +  35*3  parts  of  carbon  dioxide,  contain- 
ing 9*9  carbon,  and  25*4  oxygen  4- 2*5  parts  of  acetic 
acid,  containing  o*6  carbon,  0*2  hydrogen,  and  17 
oxygen. 


4 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.  21 


Thus  we  find  : — 

Carbon  of  the  sugar  .  ,  ,  .  26*8 
Hydrogen  „  „  .  .  .  .77 
Oxygen        >^     j?    •  • 

Sum  of  carbon  of  the  three  products  27*2 
„    „  hydrogen       „  „  9*8 

„    „  oxygen  „  „  58-5 

Taking  into  consideration  the  imperfections  of  his 
method  of  analysis,  Lavoisier  thought  the  agreement 
between  the  two  sides  of  this  equation  sufficient  to 
confirm  the  general  principle  announced  above. 

If  we  compare  with  these  numbers  those  furnished 
by  the  wonderfully  accurate  methods  employed  by 
modern  chemists,  we  shall  see  that,  in  reality,  95*9  parts 
of  cane  sugar  contain  : — 

44'4  carbon,  &i  hydrogen,  and  49*4  oxygen; 
and  give, 

51*6  of  alcohol,  containing — 

26*9  carbon,  67  hydrogen  and  i8*o  oxygen  + 

49'4  parts  of  carbon  dioxide,  containing — 

13*5  carbon,  and  36*9  oxygen. 

It  was  therefore  only  by  compensation  of  consider- 
able errors  that  Lavoisier  was  led  to  an  approximate 
solution. 

Towards  1815,  the  analyses  so  carefully  made  by 
Gay-Lussac  and  Thenard,  and  by  De  Saussure,  settled 
in  a  determinate  manner  the  composition  of  sugar  and 
of  alcohol.  These  results,  far  from  invalidating  the 
conclusions  of  Lavoisier,  gave  them  solid  support. 
Thus  Gay-Lussac  (Ann.  de  Chimie,  vol.  95,  p.  318) 
wrote :     If  it  be  supposed,  now,  that  the  products 


22 


ON  FERMENTATION. 


furnished  by  the  ferment  can  be  neglected,  as  far  as 
relates  to  the  alcohol  and  carbonic  acid  which  are  the 
only  sensible  results  of  fermentation,  it  will  be  found 
that,  given  lOO  parts  of  sugar,  51*34  of  them  will  be 
converted  during  fermentation  into  alcohol,  and  48*66 
into  carbonic  acid." 

These  results,  expressed  in  a  chemical  equation,* 
give  to  cane  sugar  the  formula  C-^^^  ^24  Oi2'i" 
we  shall  have  C^^  ^24  ^12  =  4  Hg  Og  +  4  COg.J 
The  analyses  of  cane  sugar  made  by  Gay-Lussac  and 
Thenard  themselves  agree,  as  well  as  those  since  made 
by  a   great   number  of  chemists,  with  the  formula 

In  order  to  arrive  at  the  error  which  we  have  just 
pointed  out,  and  which  Messrs.  Dumas  and  BouUay 
showed  in  1828,  Gay-Lussac  supposed  that  his  analyses 
of  cane  sugar  were  imperfect,  and  he  modified  them 
recklessly,  in  the  proportion  of  2  or  3  per  cent,  in  order 
to  establish  an  agreement  between  the  two  sides  of  his 
equation. 

"The  theory  of  fermentation  arrived  at  by  Gay- 
Lussac  is  still  imperfect,"  said  Messrs.  Dumas  and 
Boullay,  "but  it  is  no  longer  so  when  we  substitute 
ether  for  alcohol  in  the  theoretical  composition  of  sugar. 
The  most  complete  agreement  is  then  established 
between  theory  and  experiment."  The  conclusion 
which  these  two  chemists  deduced  from  this  obser- 

•  These  formulae  are  given  by  M.  Schiitzenberger  as  original  formulae.  It  may 
be  worth  while,  from  an  historical  point  of  view,  to  preserve  them  as  written  by 
their  enunciators,  in  the  '*old  notation,"  thus  : — 

t  C,2  H,.,  0,2.      I  C12  H,2  0,2  =  2  C4  Hb  O,  +  4  CO,. 
§  Cvz  H„  Oi,. 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.  23 

vation,  was  that  cane  sugar  cannot  ferment  without 
assimilating  the  elements  of  a  molecule  of  water.  In 
other  words,  Gay-Lussac's  equation,  as  a  numerical 
expression,  is  correct,  but  that  it  would  be  better  to 
write  the  first  member  of  it  under  the  form — 

C12  H22  Ou  +  H2  O  =  4  C2  He  O  +  4  CO2  * 

cane  sugar        water  alcohol      carbon  dioxide. 

A  little  later  (1832),  Dubrunfant  observed  that  before 
fermentation  commenced,  the  cane  sugar  is  transformed 
into  uncrystallizable  sugar. 

M.  Berthelot  proved  afterwards  that  the  taking  up  of 
water  by  the  cane  sugar  which  precedes  alcoholic  fer- 
mentation is  due  to  the  presence  of  a  soluble  ferment 
in  the  yeast ;  we  shall  return  again  to  this  important 
point.  Finally,  in  1833,  Biot  discovered  the  change  of 
sugar  under  the  influence  of  acids. 

Gay-Lussac's  equation,  modified  by  Dumas  and 
Boullay,  was  generally  admitted  for  more  than  twenty 
years,  as  the  mathematical  expression  of  the  decom- 
position of  sugar  by  yeast. 

Meanwhile,  in  1856,  Dubrunfant,  by  making  a  quan- 
titative analysis  of  the  carbon  dioxide  disengaged  by 
fermentation,  observed  that  it  was  not  possible  to  make 
experimentally  the  equation  of  fermentable  sugars  with 
alcohol  and  carbon  dioxide  only.  (Comp.  Rend,  de 
TAcad.  des  Sciences,  vol.  42,  p.  945.) 

The  latest  important  work  on  the  qualitative  and 
quantitative  analyses  of  the  products  of  the  alcoholic 
fermentation  of  sugars  is  due  to  M.  Pasteur.  (Ann. 


*  C12  H,i  Oil  -f  HO  =  2  C4  H,  O2  +  4  CO2. 

cane  sugar      water  alcohol      carbon  dioxide. 


24 


ON  FERMENTATION. 


Chimie  et  Physique,  3rd  series,  vol.  58,  p.  330.)  By  a 
series  of  very  interesting  researches,  and  by  irrefutable 
experiments,  this  illustrious  chemist  proves  :  1st.  That 
in  every  alcoholic  fermentation,  besides  the  principal 
products,  alcohol  and  carbon  dioxide,  glycerin  and 
succinic  acid  are  formed ;  2nd.  That  the  glycerin  and 
succinic  acid  are  produced  at  the  expense  of  the  ele- 
ments of  the  sugar,  and  that  the  ferment  takes  no  part 
in  it ;  3rd.  That,  besides  this,  the  sugar  yields  a  certain 
portion  of  its  substance  to  the  new  ferment  which  is 
developed  ;  we  shall  return  to  the  last  point  when  we 
more  especially  consider  the  ferment ;  4th.  That  the 
lactic  acid,  the  production  of  which,  in  variable  quan- 
tities, has  been  observed  in  alcoholic  fermentation,  is 
the  result  of  a  special  fermentation,  differing  from 
alcoholic  fermentation,  and  proceeding  simultaneously 
with  it. 

Let  us  say,  in  conclusion,  in  order  to  give  to  every 
one  his  due,  that  the  presence  of  succinic  acid  in  fer- 
mented liquids  had  been  observed,  before  M.  Pasteur, 
by  Dr.  Schmidt  of  Dorpat  (Handworterbuch  der  Chimie, 
Von  Liebig,  Poggendorff,  ist  edit.,  vol.  3,  p.  224,  1848), 
and  also  by  Schunck  in  the  fermentation  of  sugar 
by  means  of  erythrozyme,  the  ferment  derived  from 
madder. 

These  facts  had  passed  unperceived,  and  had  been 
forgotten,  at  the  time  when  Pasteur  returned  to  the 
study  of  this  subject. 

Without  entering  into  the  details  of  the  experiments 
on  which  Pasteur's  conclusions  rest,  and  which  will  be 
found  in  his  memoir  (loco  citato),  wc  will  simply  give 
the  results  of  his  quantitative  researches. 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.  25 

100  parts  of  cane  sugar  O^^,*  corresponding 

with  105-26  grape  sugar  2  Cg  H^^  0^.,•f  give  nearly, — 

Alcohol       .  .51*11 

Carbon  dioxide   I  4^ ^9  according  to  Gay-Lussac's  equation 

(  0*53  excess  over  „  „ 

Succinic  acid      .  0*67 
Glycerin      .  .3*16 
Matter     united)  ^.^^ 
with  ferments  > 


1 00 'GO 


Thus,  out  of  100  parts  of  cane  sugar,  about  95  parts 
are  decomposed,  according  to  Gay-Lussac's  equation  ; 
4  parts  disappear  and  form  succinic  acid,  glycerin,  and 
carbon  dioxide,  and  I  part  is  added  to  the  newly- 
formed  ferment. 

Pasteur  endeavours  to  represent  by  an  equation  the 
decomposition  of  the  4  parts  of  sugar,  which  yield 
succinic  acid  and  glycerin.  This  expression  is  very 
complex  : — 

49  (C12       On  +  H2  O) 
or  49  (C12       O12)  +  60  H2  O  =  24  (C4  H,  O4)  +  144  (C3  Hg  O3)  + 

succinic  acid  glycerin 

60  CO2.I 

carbon  dioxide 

This  equation  can  only  be  considered,  as  Pasteur 
himself  says,  as  a  very  approximate  expression  of  the 
numerical  results  of  the  analysis,  and  not  as  a  mathe- 
matical expression  of  the  reaction. 

*  C12  Hn  On.  }  C12  O12. 

1  49  (C12  Hn  On  +  HO),  or  49  (C12  iii2         +  60  HO 
12  (C,  Hg  Os)  +  72  (Co  Hg  Oo)  +  60  CO2. 


26 


ON  FERMENTATION. 


M.  Monoyer  (These  de  la  Faculte  de  Medicine  de 
Strasbourg)  proposes  a  much  more  simple  equation  to 
represent  Pasteur's  analysis  : — 

4  (Ci2       On  +  H2  O)* 
or  4  (C12  H.,  di.)  +  6  H2  O  =  2       He  O^)  +  12  (C3      O,)  ^ 
4CO2  +  O2. 

He  supposes,  at  the  same  time,  that  the  excess  of 
oxygen  serves  for  the  respiration  of  the  globules  of  the 
ferment,  a  very  plausible  interpretation,  as  we  shall 
presently  see. 

In  order  to  understand  the  chemical  possibility  of  the 
production  of  glycerin  and  succinic  acid  at  the  expense 
of  sugar,  it  is  sufficient  to  remark  that  by  adding  to- 
gether the  formulae  of  glycerin  and  succinic  acid,  atom 
by  atom,  we  arrive  at  a  sum  in  which  hydrogen  and 
oxygen  are  in  the  proportions  to  form  water : — 

C4  Hg  O4  +  C3  Hg  O3  =  C7  O7. 

On  the  other  side  we  have, 

Q       O7  +  H,  O  =  2  C3  Hg  O3  +  C  O2. 

These  two  equations  explain  naturally  enough  the 
formation  of  glycerin  and  succinic  acid,  at  the  expense 
of  glucose. 

Even  according  to  the  researches  of  Pasteur,  the  pro- 
portions of  glycerin  and  succinic  acid,  in  relation  to  the 
alcohol,  fur;iished  by  the  same  weight  of  sugar,  are  not 
absolutely  constant.  More  glycerin  and  succinic  acid, 
and  less  alcohol  are  formed,  according  as  the  fermenta- 

*  4  (C12  Hn  +  HO),  or  4  (Q2  H12  0,2)  +  6  HO  = 
Ca  H,  0^  +  6  C,  Hg  O,  +  2  C2  O,  +  O2. 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.  2/ 


tion  is  slower,  or  is  made  with  more  exhausted  and  less 
pure  yeast,  supphed  with  but  few  ahmentary  principles, 
and  those  unsuited  for  the  multiplication  of  its  globules. 

Fermentation  effected  by  sowing  the  ferment,  in  the 
presence  of  more  than  a  sufficient  quantity  of  albumi- 
noid and  mineral  matter  suited  to  the  nature  of  the 
globules,  furnishes  less  glycerin  and  succinic  acid,  and 
more  alcohol. 

A  feeble  acidity  of  the  liquor  seems  also  to  diminish 
the  proportion  of  the  two  secondary  products  ;  the  con- 
trary occurs  if  the  medium  be  neutral.  Yet  Pasteur 
himself  says,  that  we  usually  find  in  wines  a  very  large 
proportion  of  glycerin  and  succinic  acid,  although  the 
fermentation  of  the  must  of  the  grape  takes  place  in 
an  acid  medium,  in  presence  of  a  sufficient  quantity  of 
albuminoid  and  mineral  matters. 

Whatever  these  variations  may  be,  it  is  not  less  true 
that  glycerin  and  succinic  acid  were  never  deficient  in 
more  than  a  hundred  analyses  of  fermentation  made 
by  Pasteur. 

The  following  is  the  method  followed  by  M.  Pasteur, 
to  detect  and  measure  quantitatively  the  glycerin  and 
succinic  acid  contained  in  a  fermented  liquid.  The 
liquid,  when  the  fermentation  is  over,  and  all  the  sugar 
has  disappeared  (which  requires  from  fifteen  to  twenty 
days  under  good  conditions),  is  passed  through  a  filter, 
accurately  weighed  against  another  made  of  the  same 
paper.  After  having  been  dried  at  ioo°  C.  (212°  F.),  the 
dried  deposit  of  the  ferment  which  is  collected  at  the 
bottom  of  the  vessel  is  accurately  weighed.  The  filtered 
liquid  is  subjected  to  a  very  slow  evaporation  (at  the 
rate  of  from  12  to  20  hours  for  each  half-litre). 


1 
1 


28 


ON  FERMENTATION. 


When  it  is  reduced  to  loor  20  cubic  centimetres  ('61  or 
1*22  cub.  in.),  the  evaporation  is  finished  in  a  dry  vacuum. 
The  sirupy  residuum  in  the  capsule  is  treated  several  times 
with  a  mixture  of  alcohol  and  ether,  formed  of  one  part 
of  alcohol  at  30°  or  32°  and  parts  of  rectified  ether. 
After  six  or  seven  washings,  there  remains  no  more 
succinic  acid  or  glycerin.  The  etherized  alcoholic  liquid 
is  distilled  in  a  retort,  then  evaporated  in  a  water-bath 
in  a  capsule,  and  afterwards  in  a  dry  vacuum.  Pure 
lime  water  is  added  to  the  remainder,  till  it  is  neutral- 
ized ;  it  is  then  evaporated  afresh,  and  the  dried  mass 
is  again  treated  with  the  mixture  of  alcohol  and  ether, 
which  only  dissolves  out  the  glycerin,  leaving  the 
calcium  succinate  in  the  form  of  a  crystallized  powder, 
stained  with  a  small  quantity  of  extractive  matter,  and 
with  an  uncrystallizable  salt  of  lime.  The  calcium 
succinate  is  easily  purified  by  treating  it  with  alcohol 
at  80  per  cent,  which  only  dissolves  out  the  foreign 
matters.  The  etherized  alcoholic  solution  of  glycerin 
is  evaporated,  and  weighed  after  dessication,  in  a  dry 
vacuum. 

Among  the  products  met  with  normally  and  con- 
stantly in  all  alcoholic  fermentations  of  sugar,  we  ought  to 
mention  acetic  acid.  The  formation  of  this  body  noticed 
by  Bechamp  (Comp.  Rend,  de  I'Acad.,  1863),  was  at  first 
attributed  by  Pasteur  to  a  concomitant  or  subsequent 
acetic  fermentation,  and  to  the  presence  of  Mycoderma 
aceti ;  but  since  the  very  precise  and  conclusive  re- 
searches of  Duclaux  (Theses  Presentees  a  la  Faculte 
des  Sciences,  1865),  it  has  been  established:  1st.  That 
the  acetic  acid  is  never  deficient,  even  in  fermentations 
conducted  in  the  most  careful  manner,  in  order  to  pre- 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.  2Cj 

serve  them  from  contact  with  air ;  2ndly.  That  the 
proportion  of  this  acid  is  remarkably  constant,  especially 
if  we  are  careful  to  stop  the  fermentation  as  soon  as  all 
the  sugar  is  transformed ;  besides  this,  it  does  not  ex- 
ceed '05  per  cent,  of  the  weight  of  the  sugar.  This 
proportion  of  acetic  acid  is  considerably  augmented  if 
the  fermentation  be  continued  beyond  the  limits  just  indi- 
cated. As  we  shall  presently  see  that  the  ferment,  when 
left  to  itself,  without  sugar  and  without  oxygen,  can 
form  acetic  acid,  by  acting  on  its  own  elements,  we  can 
understand  the  observed  augmentation  in  the  weight  of 
this  acid,  and  we  shall  be  inclined  to  attribute  its  pro- 
duction in  a  general  manner  to  the  transformations 
undergone  by  the  ferment  while  it  acts  upon  the  sugar. 
We  may  say  the  same  thing  of  the  leucine  and  tyrosine 
found  by  Bechamp  in  the  extract  of  fermented  glucose. 
These  are  compounds  in  the  production  of  which  the 
sugar  and  the  fermentable  matter  take  no  part. 

However,  the  later  works  of  Bechamp  on  this  question 
are  not  favourable  to  this  opinion  concerning  the  acetic 
acid  (Comp.  Rend.,  vol.  75,  p.  1036).  They  tend  to 
establish:  1st.  That  the  contact  of  air,  far  from  aug- 
menting the  production  of  acetic  acid,  diminishes  it. 
A  fermentation  which  in  carbon  dioxide  produces  from 
•25  to  '40  of  this  body  for  100  of  sugar,  gives  only  *!  in 
contact  with  air  ;  2nd.  That  the  acetic  acid  comes  from 
the  sugar,  and  not  from  the  ferment,  for  by  arranging  the 
experiments  suitably,  we  may  obtain  a  weight  of  acid 
greater  than  that  of  the  ferment  employed.  In  pro- 
portion as  the  ferment  is  well  nourished,  and  the  better 
it  multiplies,  the  less  acetic  acid  it  yields.  That  which 
has  only  cane  sugar  for  its  nourishment  is  exhausted^ 


30 


ON  FERMENTATION. 


and  produces  more  acetic  acid.  The  temperature  and 
the  augmentation  of  pressure  tend  to  increase  this 
phenomenon. 

Most  of  the  natural  saccharine  juices,  such  as  those 
of  beet-root  and  grape-cake,  give  rise  to  the  production 
of  small  quantities  of  alcohols  homologous  with  ordinary 
alcohol.  We  find,  in  fact,  when  large  masses  of  pro- 
ducts are  operated  upon,  in  the  arts,  and  crude  alcohol 
is  distilled  carefully,  by  means  of  suitable  rectifying 
apparatus,  that  there  is  a  residuum  less  volatile  than 
ethyl  alcohol,  having  a  strong  and  disagreeable  odour. 
This  oily  residuum,  known  under  the  name  of  oil  of 
potatoes,  has  been  made  the  object  of  many  researches 
by  Chancel,  Wurtz,  Pelletan,  Faget,  and  others ;  they 
have  generally  found  it  composed  in  great  part  of 
propyl  alcohol,  C3  Hg  O,  butyl  alcohol  Q  H^^  O, 
dominant  amyl  alcohol,  Cg  H^^g  O,  caproic  alcohol, 
Cq  O,  oenanthyl  alcohol  C7  H^q  O,  and  caprylic 
alcohol  Cg  Hjg  O."^ 

M.  Jeanjean  has,  besides,  ascertained,  in  the  products 
of  the  distillation  of  the  water  in  which  fermented 
madder  has  been  washed,  the  presence  of  camphyl 
alcohol  (camphor  of  Borneo,  C^q  H^q  O). 

We  may  ask  whether  these  secondary  products, 
which  are  relatively  not  very  abundant,  owe  their 
origin  to  alcoholic  fermentation  properly  so  called,  or 
to  distinct  concomitant  fermentations,  having  each  a 
special  ferment  ;  or  whether,  in  fact,  it  is  better  to 
attribute  their  appearance  to  special  principles  accom- 
panying glucose  in  the  natural  saccharine  juices. 

♦  I  have  followed  Schorlcmmer  in  using  the  terms,  cthy/  alcohol,  propy/ 
alcohol,  instead  of  ethylic.    The  change  is  unimportant. 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.     3 1 


The  actual  state  of  science  does  not  allow  us  as  yet 
to  answer  these  questions  definitively. 

M.  Berthelot  points  out  (Ch.  Org.  Fondee  sur  la  Syn- 
these,  vol.  2,  p.  631)  that  the  production  of  all  these 
homologues  of  ordinary  alcohol  at  the  expense  of  sugar 
may  be  formulated  by  the  general  equation  :— 

5  I  O^  I         H^-"  +  02 

n  n— 2 

+  2^0^  +  —  H2  O. 


Of  the  Fermentable  Body. 

The  progress  of  chemistry  has  taught  us  to  distin- 
guish many  varieties  of  saccharine  hydro-carbons, 
differing  either  in  their  properties  or  in  their  compo- 
sition ;  they  do  not  all  show  the  same  characters,  when 
they  are  subjected  to  the  influence  of  the  special 
alcoholic  ferment,  the  yeast  of  beer. 

Glucose  (grape  sugar  or  starch  sugar)  levulose  or 
the  sugar  of  acid  fruits,  uncrystallizable  sugar,  maltose 
or  sugar  of  malt,  formed  by  the  action  of  diastase  on 
dextrin,  lactose  or  sugar  derived  from  sugar  of  milk 
(lactive)  by  the  action  of  acids,  all  have  the  same 
formula,  Cq  Og. 

An  almost  entire  resemblance  between  their  be- 
haviour in  presence  of  a  ferment  corresponds  with  this 
analogy  in  their  composition.  These  sugars  are  split  up 
progressively  into  alcohol  and  carbon  dioxide  without 
undergoing  any  previous  transformation- — as  Mitscher- 
lich  has  observed  (Ann.  de  Poggen.,  vol.  135,  p.  95),  the 


32 


ON  FERMENTATION. 


rotatory  power  of  a  solution  of  glucose  diminishes  in 
proportion  to  the  quantity  of  alcohol  produced. 

The  equation  of  Dumas  and  Gay-Lussac,  modified  by 
Pasteur,  applies  without  any  restriction  to  these  various 
saccharine  matters.  We  will  only  add  that,  according 
to  the  interesting  observations  of  Dubrunfant,  glucose 
mixed  with  levulose,  with  the  addition  of  yeast,  ferments 
sooner  than  the  latter  does  when  by  itself.  This  is 
what  always  happens  when,  having  altered  cane  sugar 
by  an  acid,  we  subject  to  fermentation  the  mixture  of 
equal  weights  of  glucose  and  levulose  which  results 
from  this  alteration  ;  the  glucose  disappears  before  the 
levulose,  which,  last  of  all,  undergoes  alcoholic  decom- 
position. Dubrunfant  has  given  this  phenomenon  the 
name  of  elective  fermentation.  The  sugars  whose 
composition  is  represented  by  the  formula  C^g  On 
can  also  be  fermented,  but  only  on  the  condition  of 
being  previously  hydrated,  which  converts  them  into 
sugar  with  the  formula  Cq  H^g  Oe- 

Saccharose  or  cane  sugar  is  changed,  when  hydrated, 
into  two  isomeric  molecules,  one  of  which  crystallizes 
and  causes  the  plane  of  polarized  light  to  deviate  to 
the  right,  and  the  other  remains  uncrystallizable,  and 
turns  it  to  the  left  (levulose).  The  two  products  of 
this  splitting  up  are  fermentable,  the  hydratation,  as 
is  well  known  is  effected  under  the  influence  of  acids, 
of  water  only,  of  light,  and  of  the  lower  cellular  plants. 
It  may  be  understood,  from  this  last  observation,  why 
cane  sugar  can  ferment  :  as  soon  as  it  is  placed  in 
contact  with  yeast,  it  begins  to  alter,  and  afterwards 
the  glucoses  produced  form  alcohol.  The  ferment 
therefore  plays  a  double  part  towards  the  saccharine 


ALCOHOLIC  OR  SPIRITUOUS  FERMENTATION.  33 

matter,  of  which  the  first  is  much  simpler  than  the 
second.  The  alterative  power  is  due  to  the  presence  in 
the  ferment  of  a  soluble  and  inorganic  nitrogenous 
principle,  formed  at  the  expense  of  the  proteids  of  this 
organism.  This  active  alterative  substance  accumulates 
more  especially  in  great  proportion  in  ferment  which 
has  been  left  to  itself,  and  which  has  undergone  the 
phenomenon  called  softening. 

The  water  in  which  such  ferment  has  been  washed 
alters  cane  sugar  with  such  rapidity  that,  when  we 
nix  the  two  liquids  (sweetened  water  and  the  water 
in  which  the  ferment  was  washed),  the  liquid  rapidly 
reduces  Fehling's  liquid  poured  into  it  some  seconds 
after.  We  shall  return  to  this  order  of  phenomena 
when  we  speak  of  indirect  fermentation,  called  ferment- 
ation by  a  soluble  ferment. 

Meletizose,  melitose,  and  lactine  or  sugar  of  milk, 
are  in  the  same  category  as  cane  sugar,  and  must  be 
previously  hydrated.  Berthelot  observed  this  remark- 
able peculiarity  in  melitose ;  only  half  of  this  sugar  is 
decomposed  into  alcohol  and  carbon  dioxide,  the  other 
is  transformed  into  a  compound,  isomeric  with  glucose, 
namely,  eucalin,  which  is  not  fermentable. 

All  bodies  capable  of  producing  glucose  and  its 
congeners  by  hydratation,  belong  to  the  class  of 
indirectly  fermentable  substances,  such  as  starch, 
dextrine  gum,  glycogea,  and  the  various  glucosides 
which  are  found  in  vegetable  tissues. 


34 


ON  FERMENTATION, 


CHAPTER  III. 

ALCOHOLIC  FERMENTS.^ 

We  have  hitherto  considered  bodies  susceptible  of 
alcohoHc  fermentation,  and  the  details  of  the  reaction 
known  by  this  name  ;  there  remains  for  us  to  speak  of 
the  most  interesting  part  of  our  subject,  the  exciting 
cause  of  fermentation.  It  is  more  especially  on  this, 
the  most  obscure  and  the  most  difficult  part  of  the 
question,  that  the  most  varied,  and,  we  may  say,  the 
most  lively  discussions  have  been  raised.  The  other 
parts  of  the  problem  required  for  their  solution,  in  fact, 
only  good  analysis  and  rigorous  quantitative  experi- 
ments. 

Historical, — Leuwenhoeck  was  the  first,  in  1680,  to 
examine  beer-yeast  by  the  microscope,  and  to  ascertain 
that  it  is  formed  of  very  small  spherical  or  ovoid 
globules.  He  could  not,  however,  determine  their 
nature. 

In  his  memoir  on  fermentation,  presented  to  the 
Academy  of  Florence  (1787),  Fabroni  compared  fer- 
ments to  animal  substances.  The  matter  which 
decomposes  sugar  is  a  vegeto-animal  substance  ;  it 
resides  in  peculiar  utricles,  in  grapes  as  well  as  in  corn. 

«  Cfr.— Pasteur,  "  Annales  de  Chimie  et  de  Physique,"  3rdseries,  vol.  58,  p. 
364;  Monoycr,  "  Those  de  Mcdccine,"  Strasburg,  1862;  L.  Engcl,  "Th^se 
pour  le  Doctoral  es  Sciences,"  Paris,  1872. 


ALCOHOLIC  FERMENTS. 


35 


When  the  grapes  are  crushed,  this  glutinous  matter 
is  mixed  with  the  sugar  ;  directly  the  two  substances 
come  in  contact,  effervescence  and  fermentation  com- 
mence." 

The  experiments  and  conclusions  of  Fabroni  did  not 
seem  sufficiently  to  elucidate  the  question,  for  in  the 
year  VIII.  the  class  of  physical  sciences  of  the  insti- 
tute proposed  as  the  subject  for  the  prize  the  following 
question : — 

"  What  are  the  characteristics  which,  in  vegetable  and 
animal  matters,  distinguish  those  which  serve  as  fer- 
ments from  those  which  they  cause  to  undergo  fermen- 
tation }  "  Three  years  afterwards,*  Thenard  presented  a 
remarkable  memoir  on  alcoholic  fermentation  and  fer- 
ments. He  arrived  at  the  conclusion  that  all  natural 
juices,  when  spontaneous  fermentation  has  set  up,  give 
a  deposit  which  has  the  appearance  of  beer-yeast,  and 
like  it,  is  able  to  ferment  pure  sweetened  water.  This 
ferment  is  of  an  animal  nature ;  it  is  nitrogenous,  and 
gives  over  much  ammonia  when  distilled.  When 
Thenard  said  that  ferment  is  of  an  animal  nature,  he 
considered  only  its  chemical  composition,  and  made  no 
allusion  to  the  organization  of  the  ferment.  We  shall 
return  to  the  labours  of  this  investigator  when  we 
come  to  study  the  transformations  undergone  by  the 
ferment  during  the  act  of  fermentation. 

Gay-Lussac  proves,  by  well-known  experiments,  that 
fermentation  is  only  developed  in  the  must  of  grapes 
when  it  has  been  placed  for  a  moment  in  contact  with 
air ;  he  concludes,  from  his  experiments,  that  oxygen 


"Ann.  de  Chimie,"  vol.  26,  p.  247 


36 


ON  FERMENTATION. 


is  necessary  to  commence  the  fermentation  ;  but  that 
it  is  not  required  to  continue  it 

In  1828,  Colin  made  many  experiments  which  seem 
to  show  that  a  great  number  of  organic  nitrogenous 
substances  differing  from  yeast,  and  in  process  of 
change,  are  able,  when  placed  in  sweetened  water,  to 
set  up  alcoholic  fermentation  at  the  end  of  some  hours  ; 
at  the  same  time  the  foetid  odour  of  putrefaction  is 
changed  into  the  agreeable  smell  of  the  must  of  wine 
(Colin,  Ann.  Chim.  Phys.,  2nd  series,  vol.  28,  p.  128, 
1828). 

The  question  of  fermentation  had  reached  this  point, 
and  yeast  was  regarded  as  an  immediate  principle 
of  plants,  having  the  property  of  becoming  precipi- 
tated in  presence  of  fermentable  sugars,  when  Cag- 
niard  de  Latour  took  up  the  incomplete  microscopical 
observations  of  Leuwenhoeck,  which  had  been  so  long 
forgotten. 

He  observed  that  yeast  is  a  mass  of  organic  globules, 
susceptible  of  reproducing  themselves  by  means  of 
buds,  or  seminules,  which  appeared  to  belong  to  the 
vegetable  kingdom,  and  not  to  be  simply  organic  or 
chemical  matter,  as  had  been  supposed.  He  con- 
cluded that  it  is  very  probably  by  some  effect  of  their 
vegetation  that  the  globules  of  yeast  disengage  car- 
bon dioxide  from  the  saccharine  liquid,  and  convert 
it  into  spirituous  liquon  (Ann.  Chim.  Phys.,  2nd 
series,  vol.  68.) 

The  discovery  of  Cagniard  de  Latour  was  again 
made  almost  at  the  same  time,  but  independently, 
by  Dr.  Schwann  at  Jena,  and  by  Klitzing  at  Berlin 
(Schwann,  Poggcn.  Ann.,  1837,  vol.  41,  p.  184;  Kut- 


ALCOHOLIC  FERMENTS. 


37 


zing,  Journ.  fur  Prakt  Chem.  2,  p.  385);  confirmed 
by  the  observations  of  Quevenne  (Journal  de  Pharm. 
2,  vol.  24),  of  Turpin  (Comp.  Rend,  de  TAcad.,  4,  p. 
369),  of  Mitserlich  (Poggen.  Ann.,  55,  p.  225),  it  led, 
without  any  possible  contradiction,  to  the  following 
conclusions  respecting  the  nature  of  yeast.  This  body 
was  considered  to  be  a  mass  of  organized  and  living 
cells,  composed,  like  vegetable  or  animal  cells,  of  an 
envelope  and  granular  contents. 

From  the  very  commencement,  it  was  not  agreed 
what  place  should  be  assigned  to  this  new  form  of  life. 
Some  saw  in  it  a  fungus  without  a  mycelium,  others 
looked  upon  it  as  one  of  the  algae. 

Thus  Turpin  (Comp.  Rend.,  p.  379,  1838)  placed 
the  cells  of  yeast  in  the  genus  Torula  of  Persoon,  in 
which  sporse  in  floccos  moniliformes  concatenatae,  dein 
seudentes."  These  cells  were  thus  compared  to  spores, 
without  considering  their  mode  of  production,  which  is 
quite  different,  and  without  remarking  that  the  Torula 
has  a  mycelium,  which  is  never  found  in  ferments. 

The  discovery  of  true  spores  has  since  proved  that 
ferments  cannot  be  placed  in  the  family  of  the  Toru- 
laceae.  Meyen  (Pflanzen  Physiologic,  vol.  3,  p.  455) 
also  considered  that  yeast  was  a  fungus,  and  created  a 
new  genus  for  it,  under  the  name  of  Saccharomyces. 
This  name  was  adopted  by  Rees,  Engel,  &c. ;  Kiit- 
zing,  on  the  contrary,  with  many  other  authors,  main- 
tained that  ferments  are  algae,  which  he  arranged  in  a 
separate  genus,  crypto-coccus. 

The  opinion  of  men  of  science  who  wish  to  assimi- 
late yeast  and  ferments  in  general  to  algx,  was  founded 
on  the  observation  that  these  cells  multiplied  by  budding 


38 


ON  FERMENTATION. 


only.  But  we  shall  soon  see  that  by  placing  these  fer- 
ments under  certain  conditions,  we  succeed  in  causing 
them  to  fructify  ;  besides  this,  algae  almost  always  con- 
tain chlorophyll,  which  fungi  and  ferments  do  not. 

It  is  now,  therefore,  very  generally  admitted  that 
ferments  are  fungi.  Without  in  the  least  detracting 
from  the  merits  of  Cagniard  de  Latour,  we  ought  to 
say  that  he  had  been  anticipated  in  the  field  of  micro- 
scopic observations,  not  only  by  Leuwenhoeck,  but 
also  by  Kieser  (1814,  Schweigger's  Journal,  12,  p.  229), 
who  describes  it  as  formed  of  small  transparent  motion- 
less spherical  corpuscules,  all  of  nearly  the  same  size  ; 
also  by  Desmazieres,  who,  in  1826  (Annales  des  Sciences 
Naturelles,  vol.  10,  p.  4),  examined  the  pellicle  formed 
on  the  surface  of  beer,  and  called  by  Persoon  viyco- 
denna  cerevisice, 

Desmazieres  was  the  first  to  give  a  representation  of 
the  globules  which  he  had  observed^  and  having  seen 
in  them  a  particular  movement,  which  is  nothing  more 
than  the  Brownian  movement,  which  at  that  time  was 
unknown,  he  arranged  these  globules  among  the  ani- 
malcula  monadina.  Astier,  as  early  as  181 3  (Ann.  de 
Chimie,  vol.  87,  p.  271),  did  not  hesitate  to  affirm  that 
ferment,  recognized  as  an  animal  substance  by  Fabroni, 
was  alive,  and  derived  its  nourishment  from  the  sugar, 
whence  resulted  the  rupture  of  equilibrium  between  the 
elements  of  this  body.  By  this  theory,  it  is  easily  ex- 
plained, said  he,  that  all  the  causes  which  kill  animals, 
or  hinder  their  development,  must  be  opposed  to  fer- 
mentation. 

The  observers  who  had  demonstrated  the  organic 
nature  of  ferments  established  at  the  same  time  that 


ALCOHOLIC  FERMENTS. 


39 


in  a  great  number  of  liquids  in  alcoholic  fermentation 
(such  as  natural  saccharine  juices,  solutions  of  sugar 
with  albumin,  &c),  globules  of  ferment  are  formed,  as 
Thenard  had  observed.  Schmidt  of  Dorpat  arrived  at 
the  same  conclusions  by  repeating  Colin's  experiments 
on  the  fermentation  excited  by  albuminoid  matter  in 
the  process  of  decomposition.  Microscopic  observa- 
tions revealed  to  him  the  development  of  globules  of 
ferment  whenever  there  was  the  production  of  alcohol. 

From  all  these  successive  observations,  which  were 
complementary  to  each  other,  arose  the  opinion  generally 
admitted,  that  yeast  accompanies  every  well  marked 
alcoholic  fermentation ;  it  seems,  therefore,  that  the 
theory  propounded  by  Astier  and  Cagniard  de  Latour 
concerning  fermentation,  ought  to  have  prevailed,  and 
to  have  been  received  by  men  of  science.  But  this  was 
not  the  case.  From  the  very  commencement  of  the 
discussion,  the  conclusions  of  Cagniard  de  Latour  and 
of  Schwann  found  a  powerful  opponent.  Liebig,  whose 
name  was  then  an  authority  in  chemistry,  had  a  decided 
theory  concerning  the  phenomena  of  fermentation  in 
genelal,  and  he  defended  it  with  vigour,  even  after  the 
experiments  of  Pasteur,  who  admits  that  alcoholic  fer- 
mentation is  an  act  connected  with  life,  and  with  the 
organization  of  globules. 

"My  most  decided  opinion,"  says  Pasteur,  *'on  the 
nature  of  alcoholic  fermentation  is  the  following :  The 
chemical  act  of  fermentation  is  essentially  a  correlative 
phenomenon  of  a  vital  act,  beginning  and  ending  with 
it.  I  think  that  there  is  never  any  alcoholic  fermentation 
without  there  being,  at  the  same  time,  organization, 
development,  multiplication  of  globules,  or  the  con- 


40 


ON  FERMENTATION. 


tinued  consecutive  life  of  globules  already  formed." 
As  to  the  hypotheses  which  tend  to  go  more  deeply  into 
the  physiological  cause  of  decomposition,  M.  Pasteur 
neither  admits  nor  rejects  them,  at  least  in  his  first 
memoir.  Thus  the  ideas  of  Pasteur  confirm  and  extend 
those  of  Cagniard  de  Latoun 

As  to  the  theory  ofJLiebig,  it  does  not  differ  from 
that  of  Willis  and  Stahl.  According  to  the  German 
chemist,  the  cause  of  fermentation  is  the  internal 
molecular  motion,  which  a  body  in  the  course  of  de- 
composition communicates  to  other  matter  in  which 
the  elements  are  connected  by  a  very  feeble  affinity. 
"Yeast,  and,  in  general,  all  animal  and  vegetable  matters 
in  a  state  of  putrefaction,  will  communicate  to  other 
bodies  the  condition  of  decomposition  in  which  they 
are  themselves  placed  ;  the  motion  which  is  given  to 
their  own  elements  by  the  disturbance  of  equilibrium  is 
also  communicated  to  the  elements  of  the  bodies  which 
come  into  contact  with  them.  (Liebig,  Ann.  de  Chimie 
et  de  Phys.,  2nd  series,  vol.  71,  p.  178.)  This  very 
philosophical  and  seducing  explanation  of  an  obscure 
phenomenon  obtained  greater  credit  among  men  of 
science  because  it  gave  the  key,  not  only  to  alcoholic  fer- 
mentation, but  also  to  other  phenomena  of  the  same 
kind,  such  as  the  transformation  of  sugar  into  lactic 
and  butyric  acids,  in  w^hich  organic  products  had  not 
hitherto  been  observed,  and  which  seemed  only  to  be 
the  results  of  a  conflict  between  a  fermentable  substance 
and  a  nitrogenous  substance  in  process  of  putrefaction. 
]^>cmy  and  I^outron  supposed  that,  in  matters  capable 
of  acting  as  ferments,  the  character  of  the  fermentation 
varies  with  the  degree  of  decomposition  of  the  sub- 


ALCOHOLIC  FERMENTS. 


41 


stances.  This  would  be  successively  alcoholic,  lactic,  or 
butyric  ferment  according  to  the  more  or  less  advanced 
state  of  its  decomposition. 

It  is  thus  that  the  recognized  invariable  presence  of 
an  organic  body  in  every  liquid  in  process  of  alcoholic 
fermentation  began,  by  degrees,  to  be  considered  as  a 
fact  of  slight  importance  with  regard  to  the  reaction  ; 
the  latter  is  excited,  not  by  the  direct  action  of  the 
globules  of  ferment,  considered  as  a  living  organism, 
but  by  the  decomposition  of  the  proteic  nitrogenous 
matter  of  this  ferment,  regarded  only  as  nitrogenous 
substance.  The  experiment  of  Gay-Lussac  was 
naturally  interpreted  by  this  opinion  ;  the  momentary 
presence  of  oxygen  was  indispensable  to  set  up  the 
molecular  disturbance  of  the  albuminous  matter  of 
the  must  of  grapes. 

Berzelius,  for  his  part,  treated  the  organic  nature  of 
yeast  as  a  poetico-scientific  reverie,  and,  rejecting  the 
doctrine  of  Liebig,  borrowed  from  Willis  and  Stahl, 
would  only  see  in  fermentation  an  act  of  contact  due  to 
catalytic  force,  and  in  yeast  an  amorphous  principle. 
Mitscherlich  supported  the  ideas  of  Berzelius,  while  he 
admitted  the  organic  nature  of  the  ferment. 
.  However,  the  clear  and  well  conducted  researches  of 
Pasteur  on  fermentation,  and  especially  on  alcoholic 
fermentation,  had  partly  reconciled  men  of  science  to 
the  physiological  theory ;  wherefore  Liebig  thought  right 
to  recommence  the  contest  in  favour  of  his  own  ideas. 
In  1870,  he  published  a  long  m.emoir  on  fermentation 
and  the  source  of  muscular  force  (Ann.  der  Chemie 
und  Pharmacie,  vol.  153,  p.  i),  a  memoir  in  which  he 
sought  to  show  that  the  principal  experiments  of  Pasteur 
3 


42 


ON  FERMENTATION. 


are  not  conclusive.  He  first  demonstrated  that  the 
physiological  theory  of  Pasteur,  which  explains  the  de- 
composition of  sugar  by  the  nutrition  and  development 
of  an  organic  substance,  is  not  opposed  to  the  mechani- 
cal doctrine  of  which  he  is  the  champion.  This  avowal 
was  already  an  enormous  concession  made  by  the  Ger- 
man chemist,  almost  an  avowal  of  defeat,  for  this  lan- 
guage is  very  different  from  that  which  he  had  formerly 
used. 

However,  the  attack  was  sufficiently  powerful  to 
induce  Pasteur  to  reply  to  it  (Ann.  Chimie  Phys.,  4th 
series,  vol.  25,  p.  145,  1872),  and  Dumas  to  undertake 
a  series  of  experiments,  in  order  to  ascertain  if  it  were 
possible  to  verify  the  consequences  of  Liebig's  theory. 

Dumas,  by  means  of  ingenious  experiments,  con- 
ducted with  his  never-failing  precision  (Ann.  de  Chimie 
et  de  Physique,  5  th  series,  vol.  3,  p.  69),  succeeded  in 
proving  irrefutably;  1st.  That  saccharine  liquids  are 
not  influenced  by  ferment,  even  through  the  shortest 
columns  of  liquid,  the  thinnest  membranes,  or  even  with- 
out any  separating  medium  ;  and  that  its  immediate 
and  direct  contact  is  necessary ;  2nd.  That  sonorous 
vibrations  have  no  influence  on  the  movements  of 
fermentation  ;  3rd.  That  no  chemical  action,  among  a 
great  number  of  those  which  have  been  tried,  has  been 
able  to  effect  the  decomposition  of  sugar  into  alcohol 
and  carbon  dioxide. 

These  negative  results,  without  bringing  about  any 
decisive  solution  of  the  question,  are,  however,  contrary 
to  the  opinion  of  a  transmitted  movement. 

We  may  thus  sum  up  these  three  great  theories  of 
fermentation:  1st.  The  vitalist  theory,  formulated  by 


i 


ALCOHOLIC  FERMENTS. 


43 


these  words  of  Turpin,  "  Fermentation  as  effect,  and 
vegetation  as  cause,  are  two  things  inseparable  in  the 
act  of  decomposition  of  sugar,"  maintained  by  Astier, 
Cagniard  de  Latour,  Schwann,  Kiitzing,  Turpin,  Bou- 
chardot.  Van  de  Brock,  Shroeder,  Pasteur,  and  Bichat ; 
2nd.  The  mechanical  theory  of  Willis,  Stahl,  and 
Liebig,  admitted  by  Gerhardt ;  3rd.  The  theory  of 
catalytic  forces,  and  of  acts  of  contact,  maintained  by 
Berzelius  and  Mitscherlich. 

Various  mixed  opinions  range  themselves  by  the  side 
of  these  three  theories.  Thus  M.  Berthelot  considers 
fermentation  as  produced  by  the  action  of  a  substance 
elaborated  by  organic  ferments,  comparing,  with  this 
idea,  the  alcoholic  and  lactic  fermentations  to  the  con- 
version of  starch  into  dextrine  and  sugar  under  the 
influence  of  diastase,  a  soluble  inorganic  ferment.  The 
learned  chemist  supports  his  opinion  by  experiments 
which  prove  that,  in  certain  cases,  there  may  be  the 
production  of  alcohol  without  the  formation  of  ferment 
(Chimie  Organique  Fondee  sur  la  Synthese)  ;  which  does 
not  exclude  the  fact,  now  distinctly  established,  that 
fermentation  may  be  excited,  and  is  indeed  energetically 
originated,  by  special  organic  substances. 

As  to  a  more  precise  relation  between  chemical 
phenomena  and  the  physiological  functions  of  the 
organic  ferrrient,  it  is  still  to  be  discovered  ;  and  all 
that  has  been  said,  written,  and  brought  forward  to 
decide  the  question  needs  experimental  proof,  and  can 
only  be  considered  by  us  in  passing. 

No  one  doubts  that,  in  organic  living  cells,  whether 
they  be  isolated,  like  those  of  yeast,  or  form  an  integral 
part  of  a  more  complicated  organism,  there  resides  a 


44  ON  FERMENTATION. 

special  force,  capable  of  producing  chemical  reactions 
under  conditions  quite  different  from  those  which  we 
employ  in  our  laboratories,  and  to  produce  results  of  the 
same  class.  This  force,  which  we  imagine  to  be  as 
material  as  heat,  reveals  to  us  its  activity  by  decom- 
positions effected  on  complex  molecules.  Whether  we 
reduce  the  problem  to  the  action  of  a  soluble  product 
elaborated  by  the  organic  ferment,  and  to  which  it  has 
communicated  its  power,  or  suppose  that  the  whole  of 
the  ferment  exercises  an  action  of  this  kind,  we  ultimately 
arrive  at  a  motion  communicated,  more  or  less  directly, 
by  vital  force,  and  dependent  upon  it. 

We  must  not  confound  this  interpretation  of  the 
phenomena  with  Liebig's  theory  ;  the  German  chemist 
held  that  the  albuminoid  principles,  in  decomposing 
spontaneously,  produce  the  molecular  motion  which  is 
transmitted  to  the  sugar  to  split  it  up  ;  here,  on  the 
contrary,  it  is  the  living  organism  which  develops  force, 
by  borrowing  it  from  the  great  external  reservoir  and 
transforming  it.  This  force  may  act  chemically,  either 
directly  or  indirectly  by  means  of  a  soluble  ferment. 

Description  of  the  Ferme^it, — We  will  give,  with  Rees, 
the  name  Saccharomyces  cerevisice^  to  the  alcoholic 
ferment  of  beer ;  it  is  the  ferment  which  has  been 
most  thoroughly  studied,  and  which  is  the  most  easily 
procured. 

There  are  three  methods  of  causing  the  wort  of  beer 
to  ferment ;  the  two  first  are  generally  employed  ;  these 
arc  surface  and  sedimentary  fermentations. 

In  the  third  method,  employed  in  Belgium  only,  the 
wort  is  left  to  itself  in  a  locality  situated  above  the 
level  of  the  ground,  and  the  spontaneous  development 


ALCOHOLIC  FERMENTS. 


45 


of  fermentation  is  waited  for ;  in  the  two  former  in- 
stances, the  action  is  excited  by  mixing  with  the  wort 
a  suitable  proportion  of  yeast  arising  from  an  anterior 
operation  of  the  same  class. 

In  beer  brewed  by  surface  fermentation,  the  starch  of 
the  malt  is  changed  into  sugar  by  its  being  steeped 
several  times  successively ;  fermentation  takes  place  in 
casks  at  a  comparatively  high  temperature,  from  15°  to 
18°  C.  (Sg""  to  about  65°  Fahr.)  The  yeast,  in  this 
case,  as  it  is  formed,  rises  through  the  bung-holes, 
at  the  upper  part  of  the  cask.  In  England,  this  fer- 
mentation is  carried  on  in  large  open  vats ;  the  yeast 
then  floats  on  the  surface  of  the  liquor,  and  can  be 
skimmed  off". 

In  the  manufacture  of  beer  brewed  by  sedimentary 
fermentation,  the  saccharification  is  effected  by  de- 


FiG.  I.  "Saccharomyces  cercvIs'iJE— Yeast  of  sedimentary  beer,  X  400. 

coction,  and  the  transformation  takes  place  in  open 
vats,  at  a  temperature  not  exceeding  from  12°  to  14°  C. 
(from  about  53°  to  58'' Fahr.)  The  yeast  is  deposited 
at  the  bottom  of  the  vats,  and  adheres  in  the  form  of  a 
pasty  mass.  When  once  the  first  and  most  active 
fermentation  is  over  (it  usually  lasts  two  or  three  days 
for  the  surface  fermentation,  and  eight  or  ten  days  for 
the  sedimentary),  the  clear  liquid  is  drawn  off,  and  kept 


46 


ON  FERMENTATION. 


in  casks,  glass,  or  stone  bottles.  In  the  meantime  the 
separation  of  the  yeast  has  not  been  completed  ;  it 
continues  to  act  on  the  still  unmodified  sugar ;  therefore 
the  amount  of  alcohol  and  carbon  dioxide  yielded 
increases  with  the  time  of  keeping,  and  at  the  same 
time  the  liquid  becomes  turbid  by  the  production  of 
fresh  yeast. 

The  yeast  greatly  exceeds  (seven  or  eight  times),  in 
weight  and  in  volume,  that  which  had  been  previously 
introduced  into  the  wort.  To  this  fact,  which  we  now 
merely  notice  in  passing,  we  shall  return  presently  ;  it 
is  explained  by  the  multiplication  by  buds,  which  takes 
place  whenever  the  cells  of  yeast  are  placed  in  a  sac- 
charine medium  suitable  to  their  development ;  and 
the  wort  of  beer  affords  excellent  conditions  in  this 
respect.  After  the  sedimentary  fermentation,  the  yeast 
found  at  the  bottom  of  the  vat  is  composed,  almost 
entirely,  of  cells  of  a  single  species  of  alcoholic  ferment, 
the  Saccharomyces  cerevisice  (Fig.  i ).  The  mitroscope  will 
show,  but  in  a  very  small  proportion,  granules  of  lupu- 
lin,  crystals  of  calcium  oxalate,  spores  and  mould. 
This  deposit  is  of  the  consistence  of  a  paste  of  a  yellow- 
ish white,  or  yellow-ochre  colour. 

The  cells  are  round  or  oval,  from  l  oVo 

of  a  millimetre  (from  '00031  to  '00035  in.)  in  their 
greatest  diameter.  They  are  formed  of  a  thin  and 
clastic  membrane  of  colourless  cellulose,  and  of  a  pro- 
toplasm, also  colourless,  .sometimes  homogeneous,  some- 
times composed  of  small  granulations.  We  find  in  the 
protoplasm  one  or  two  vacuoles,  of  various  sizes,  con- 
taining cellular  juice.  The  cells  arc  cither  separate,  or 
united  two  by  two. 


ALCOHOLIC  FERMENTS. 


47 


When  these  cells  are  deposited  in  a  fermentable 
liquid,  we  soon  see  at  one,  and  more  rarely,  at  two  points 
of  their  surface,  vesicular  prominences  arise,  the  interior 
of  which  is  filled  at  the  expense  of  the  protoplasm  of 
the  mother-cell ;  these  prominences  enlarge,  and  at  last, 
having  attained  the  size  of  the  original  cell,  they  lessen 
in  diameter  at  the  base  (Fig.  2).  They  usually  originate 
at  the  widest  side,  but  more  rareiy  at  the  extremities. 
As  soon  as  the  formation  of  this  kind  of  neck  takes 
place,  the  new  cells  separate  with  considerable  rapidity 
from  the  mother-cell,  in  which  the  protoplasm,  given  up 
to  the  young  cell,  is  replaced  by  one  or  two  vacuoles. 
If  the  conditions  are  favourable,  the  same  cell  is  able  to 
produce  several  generations  of  cells  ;  but,  by  degrees,  it 
loses  all  its  protoplasm,  which  at  last  unites  in  granules 
swimming  in  the  midst  of  superabundant  cellular  juice. 
The  cell  then  ceases  to  reproduce,  and  even  to  live  ;  the 
membrane  is  ruptured,  and  the  granular  contents  are 
diffused  in  the  liquid. 


Fig.    2.  —  Saccharomyces    cerevislse —         Fig.    3.  —  Saccharomyces  cerevisiae — 
Yeast  of  sedimentary  beer,  budding,  X       Yeast  of  surface  beer,  budding,  x  400. 
400. 


When  the  Saccharoinyces  cerevisice  is  not  in  contact 


48 


ON  FERMENTATION. 


with  a  fermentable  liquid,  it  may  remain  for  some  time 
without  becoming  modified. 

The  isolated  cells  of  the  surface  ferment  (Fig.  3)  do 
not  differ  sensibly  from  those  of  the  sedimentary  fer- 
ment ;  and  although  it  has  been  maintained  that  the 
larger  and  oval  forms  are  more  prevalent  in  it,  it  is  diffi- 
cult to  establish  any  distinction  of  this  kind,  for  we  find 
in  the  two  varieties  all  the  intermediate  forms  between 
the  two  extremes. 

Besides,  an  elevation  of  temperature  above  14''  C. 
(57*2  Fahr.)  during  the  fermentation  is  sufficient  to 
augment  considerably  the  size  of  the  sedimentary 
cells,  to  cause  them  to  attain  a  large  diameter,  from 
liio  to  jiio  of  a  millimetre  ('00051  to  '00055  of 
an  in.),  giving  them  a  long  oval  form  ;  at  the  same 
time,  we  see  two  circular  vacuoles  make  their  ap- 
pearance ;  one,  large,  situated  near  the  larger  end ; 
the  other,  smaller,  is  found  in  the  narrow  part  of  the 
cell. 

The  surface  SaccJiaj'Oinyccs  buds  much  more  quickly 
than  the  other  variety,  when  placed  in  a  fermentable 
liquid  (Fig.  3).  This  budding  is  very  rapid  ;  the  different 
cells  which  issue  from  each  other  remain  attached 
together,  forming  small  ramified  chains,  composed  of 
from  six  to  twelve,  and  even  more,  individual  buds.  It 
may  be  easily  understood  that  the  bubbles  of  gas 
adhering  to  these  chaplets  have  greater  hold  upon  them 
than  on  an  isolated  cell  ;  this  causes  the  newly-formed 
yeast  to  be  raised  towards  the  surface  of  the  liquid,  and 
this  is  effected  the  more  rapidly  when  the  fermentation 
is  more  active.  In  these  chaplets,  the  cells  have  an 
elliptical  form.  ^The  only  well- ascertained  difference 


ALCOHOLIC  FERMENTS. 


49 


between  the  two  kinds  of  yeast  is,  therefore,  the  rapidity 
with  which  buds  are  formed,  and  the  greater  activity  of 


Fig.  4. — Saccharomyces  cerevisicje-— Sur-  Fig.  5. — Saccharomycescerevisiae— Sedi- 
face  yeast,  at  rest,  x  400.  mentary  yeast,  in  a  growing  state,  X  400. 

the  ferment  in  the  one  case ;  but  this  does  not  authorize 
us  to  consider  these  two  ferments  as  belonging  to  different 
species.  We  are  able,  indeed,  though  with  great  diffi- 
culty, by  changing  their  conditions  of  existence,  to 
transform  one  into  the  other. 

Multiplication  by  buds  is  not  the  only  mode  of  repro- 
duction of  the  Saccharomyces  cerevisice.  It  is  true  that 
it  alone  appears  as  long  as  the  yeast  is  in  contact 
with  an  appropriate  fermentable  liquid.  We  owe  to 
Rees  (Botanische  Zeitung,  December,  1869)  the  dis- 
covery of  the  fructification  of  yeast,  and  of  ferments  in 
general ;  that  is  to  say,  their  reproduction  by  means  of 
spores.  As  to  the  Saccharomyces^  the  conditions  which 
appear  to  be  peculiarly  favourable  to  this  special  evolu- 
tion of  the  fungus,  are  to  deprive  it  suddenly  of  all 
nourishment,  especially  saccharine,  and  to  expose  it 
to  a  damp  atmosphere,  or,  still  better,  to  place  it  on  a 
substance  capable  of  affording  it  sufficient  and  constant 
humidity. 

We  obtain,  according  to  Rees,  the  richest  production 
of  spores  by  leaving  yeast  previously  washed  several 
times,  for  some  days  in  contact  with  distilled  water,  and 


50 


ON  FERMENTATION. 


then,  having  decanted  the  greater  part  of  the  water, 
later  on  removing  day  by  day  the  small  portions  of 
water  which  become  separated  from  it.  Under  favour- 
able conditions,  we  obtain,  at  the  end  of  fifteen  or  six- 
teen days,  a  very  rich  formation  of  spores  ;  but  very 
often  this  result  is  prevented  by  the  putrefaction  of  the 
yeast. 

M.  Engel,  from  whose  essay  we  borrow  the  greater 
part  of  these  details,  has  also  studied  this  question :  he 
proposes  to  cause  the  yeast  to  fructify,  by  the  following 
contrivance : — 

We  mix  up  some  plaster,  and  allow  it  to  run  on  some 
polished,  but  not  oily  surface,  such  as  window-glass, 
plate-glass,  or  marble.  We  make  up  the  mass  into 
some  form  corresponding  with  the  interior  of  the  vessel 
in  which  we  intend  to  preserve  it.  The  dimensions  in 
each  direction  ought  to  be  about  two  centimetres 
(78  in.)  smaller  than  the  internal  dimensions  of  the 
vessel,  so  as  to  allow  a  space  between  the  sides  and  the 
mass  of  plaster  sufficient  to  pour  in  some  distilled 
water.  We  then  take  some  very  fresh  yeast,  decant  as 
far  as  possible  all  the  supernatant  fermentable  liquid, 
and  dilute  the  yeast  with  distilled  water,  so  as  to  bring 
it  to  the  consistence  of  very  fluid  broth  ;  we  pour  some 
drops  of  this  liquid  on  the  polished  surface  of  the 
plaster,  inclining  the  mass  in  every  direction  so  as  to 
spread  the  solution  uniformly.  This  operation  should 
be  performed  quickly,  for  the  plaster  absorbing  the 
water  very  rapidly,  the  diluted  yeast  would  become  too 
thick,  would  not  spread  with  sufficient  uniformity,  and 
the  layer  of  ferment  would  be  too  thick  in  certain  parts. 
We  then  place  the  mass  in  the  vessel,  with  the  part 


ALCOHOLIC  FERMENTS. 


51 


covered  with  yeast  upwards,  and  pour,  by  means  of  a 
funnel,  distilled  water  between  the  sides  of  the  vessel 
and  the  piece  of  plaster,  until  the  liquid  reaches  above 
a  centimetre  ('39  of  an  in.)  beneath  its  upper  surface. 
The  vessel  is  then  covered  with  a  plate  of  glass  to 
prevent,  as  far  as  possible,  the  contact  of  dust,  and  of 
spores  floating  in  the  atmosphere. 

Under  these  conditions,  the  vegetative  life  of  the  yeast 
ceases  suddenly,  and  in  a  few  hours  we  see  great 
changes  take  place  in  the  protoplasm  of  the  cells.  The 
oldest,  and  those  which  are  least  rich  in  protoplasm, 
perish  and  break  up.  Others,  on  the  contrary,  grow 
larger,  their  lacunae  disappear,  auvd  the  protoplasm  is 
diffused  uniformly  in  the  cellular  juice.  At  the  expira- 
tion of  from  six  to  ten  hours,  we  notice  the  appearance, 
in  the  midst  of  the  protoplasm,  of  from  two  to  four  small 
"  islets,"  more  brilliant  and  dense  than  the  rest,  around 
which  fine  granulations  collect.  These  dense  spots  do 
not  present  any  appearance  of  a  nucleus,  and  they 
become  differentiated  more  and  more  till  they  are 


Fig.  6. — Saccharomyces  cerevisiae — For- 
mation of  spores,  X  750.  a,  b,  c,  d,  suc- 
cessive phases  oi  the  production  of  spores. 


Fig.  7. — Triads  of  spores,  germinating, 
X  750. 


exactly  spherical  (Figs.  6  and  7).  Twelve  or  twenty- 
four  hours  later,  each  of  these  spherules  is  invested  with 
a  membrane,  very  thin  at  first,  but  which  tliickens  by 


52 


ON  FERMENTATION. 


degrees,  and  then  shows  a  double  outline,  when  magni- 
fied 600  diameters. 

The  spore  is  then  ripe.  The  mother-cell  thus  con- 
tains from  two  to  four  spores.  When  there  are  but  two, 
they  are  situated  in  the  greater  diameter  ;  when  there 
are  three,  they  are  usually  arranged  in  a  triangle ;  when 
there  are  four,  they  are  in  the  form  of  a  cross,  or  three 
of  them  form  a  triangle,  on  which  the  fourth  is  super- 
posed in  the  manner  of  a  tetrahedron. 

During  their  evolution  the  spores  touch  each  other ; 
a  plane  surface  is  therefore  produced  at  the  point  of 
contact ;  they  remain  attached  to  each  other  during 
some  time  after  maturity,  and  thus  form  combinations 
of  two,  three,  and  four.  The  two  spores  connected  to- 
gether have  only  one  plane  surface,  the  triads  have  two, 
inclined  to  each  other  at  about  120°,  and  in  the  tetrads 
arranged  in  the  form  of  a  cross  we  observe,^  also,  two 
plane  surfaces  at  right  angles.  When  the  spores  ripen, 
the  thecae  are  moulded  on  them,  and  thus  assume  their 
various  forms.  The  theca  of  the  diads  is  elliptical ; 
that  of  the  triads  is  triangular,  with  rounded  angles  ; 
that  of  the  tetrads,  in  the  shape  of  a  cross,  is  in  the  form 
of  a  diamond  with  rounded  angles  ;  in  the  tetrads,  piled 
up  on  each  other,  the  theca  is  tetrahedral ;  when  in 
complete  maturity,  the  membrance  of  the  spore  case,  or 
mother-cell,  which  has  become  a  fruit,  is  torn,  and  allows 
the  spores  to  escape.  The  thecal  themselves  vary  from 
yioo  to  xlfro  of  a  millimetre  ('00039  to  '00047  of  an  in.), 
and  the  spores  from  -y^^o^  to  tooo  of  a  millimetre  (-00015 
to  -00019  in.) 

Tlic  innumerable  quantity  of  thcc:u  which  we  obtain 
by  the  method  of  fructification  on  plaster,  leaves  no 


ALCOHOLIC  FERMENTS. 


53 


doubt  as  to  the  origin  of  these  organisms;  besides,  Rees 
and  Engel  have  often  observed  thecae  still  attached  to 
vegetative  cells,  and  those  in  process  of  budding,  and 
have  thus  recognized  their  relationship. 

Hitherto,  all  that  we  have  said  applies  to  sedimentary 
beer,  properly  so  called,  to  the  Saccharoniyces  cerevisice ; 
but  this  special  fungus  is  not  the  only  one  capable  of 
determining  the  alcoholic  fermentation  of  glucose. 
Microscopists  who  have  studied  this  difficult  question 
distinguish,  besides  the  special  ferment  of  beer,  other 
varieties,  for  the  most  part  belonging  to  the  genus  Sac- 
charomyces  of  Meyen.* 

Under  the  name  of  Saccharomyces  minor^  Engel 
describes  a  kind  of  ferment  obtained  by  him  from  the 
leaven  of  flour,  and  to  which  it  owes  its  activity. 

The  process  of  extraction  is  similar  to  that  employed 
by  chemists  to  separate  starch  from  gluten  in  flour. 
The  liquid  which  passes  through  the  sieve  when  bakers* 
yeast  is  kneaded  under  a  slight  stream  of  water,  con- 
tains starch  and  globules  of  yeast,  which,  on  account  of 
their  smaller  density,  are  deposited  last.  We  may  thus, 
by  a  series  of  washings,  obtain  a  product  very  rich  in 
globules  of  yeast,  and  poor  in  grains  of  starch.  Engel 
proposes  to  employ  sweetened  instead  of  pure  water  in 
these  washings,  in  order  not  to  diminish  the  physiologi- 
cal activity  of  the  cells. 

*  Simple  thecaphorous  fungi,  without  a  true  mycelium.  The  vegetative 
organs  are  cells,  produced  usually  by  buds  from  similar  cells,  and  which,  de- 
taching themselves,  sooner  or  later,  from  the  mother-cell,  multiply  in  the  same 
manner.  A  part  of  the  cells  thus  formed  are  transformed,  in  another  medium, 
into  naked  sporiferous  thecae  ;  unicellular  spores,  to  the  number  of  from  one 
to  four  in  each  theca.  The  germination  of  spores  reproduces  directly  vege- 
tative cells  analogous  to  those  which  originate  from  buds.— Engel,  Thesis  of 
the  Factdty  of  Sciences  of  Paris,  1872. 


54 


ON  FERMENTATION. 


The  ferment,  examined  by  the  microscope,  is  seen 
under  the  form  of  isolated  globules,  double  or  some- 
times united  in  threes.  The  largest  of  these  globules 
are  ^^^^  ^  millimetre  (about  '000315  of  an  in.)  in 
diameter ;  their  vacuoles  are  less  apparent  than  those 
of  the  yeast  of  beer. 

This  ferment,  sown  in  the  most  favourable  saccharine 
medium,  prepared  according  to  the  formula  of  M.  Pas- 
teur, has  only  produced  a  very  slow  fermciitation.  By 
renewing  the  experiment  seven  times,  and  only  making 
use  each  time  of  the  ferment  obtained  in  the  preceding 
experiment,  we  see  no  apparent  modification  in  the  form 
and  dimensions  of  the  globules.  The  budding  of  this 
species  is  effected  in  the  same  manner  as  in  the  yeast  of 
beer. 

Placed  under  the  conditions  favourable  to  the  forma- 
tion of  spores,  of  which  we  have  spoken  before,  the 
Saccharomyces  minor  is  transformed  into  sporiferous 
thecse  of  spherical,  and  occasionally  though  rarely  of 
ovoid  form,  of  yo^oo  to  y^^oo  of  a  millimetre  in  dia- 
meter. The  spores  are  only  y^^o  ^  millimetre  in 
diameter,  and  are  united  in  diads  or  triads.  In  fact, 
except  their  form,  which  is  always  spherical,  their  smaller 
dimensions  and  greater  activity,  the  ferments  of  bread 
resemble  that  of  beer. 

The  Saccharomyces  ellipsoideiis  of  Rees  is  nothing  but 
Pasteur's  ordinary  alcoholic  ferment  of  wine  (Etudes 
sur  Ic  Vin,  Figs.  8,  9,  and  ii);  it  ought  not  to 
be  confounded  with  the  Cryptococciis  vini  of  Kutzing. 
The  adult  cells  have  an  ellipsoidal  form  t/;jo  of  a  milli- 
metre in  length  by  y^,^^,^  or  y  ^  in  breadth  (-00024  by 
about  -000176  in.),  with  an  oval  vacuole.    The  sporula- 


ALCOHOLIC  FERMENTS. 


55 


Hon  and  budding  differ  in  no  respect  from  the  analogous 
phenomena  which  are  observed  in  yeast  (Figs.  8,  9, 
and  10). 

Rees  gave  the  name  of  Saccharornyces  Pastorianiis 
(Fig.  11),  to  a  variety  of  alcoholic  ferment  of  wine 
observed  by  Pasteur  (Fig.  7,  Etudes  sur  le  Vin).  The 
cells  are  oval,  pyriform,  or  elongated  like  a  club. 
The  ovoid  cells  are  -^-§-^  of  a  millimetre  in  length 
(•000236  in.),  those  that  are  club-shaped,  which  are  seen 
to  proceed  like  buds  from  ovoid  cells,  reach  y||^  or 


_  Fig.  8.— Saccharornyces  ellipsoideus,  Fig.  9.  -Saccharornyces  ellipsoideus, 
m  process  of  budding,  x  600.  development  of  spores,  X  400. 

To-oo  of  a  millimetre  in  length,  by  to^^o  or  -^1%^  in 
breadth  at  the  larger  end  ('00064  ii^-  to  -00078  in.  by 
•000314  in.  to  -000397  in.);  they  are  united  in  flakes 
containing  from  three  to  seven  articulations. 


Fig.  10, 

The  Saccharoinyces   exiguus  (Rees;,  Fig.  12,  is  met 


56  ON  FERMENTATION. 

with,  like  the  preceding,  in  the  juices  of  fermented  fruits. 
The  cells  are  only  q{  ^  millimetre  in  length,  by 
tI'Io  width  at  the  larger  end  ('0001 1 8  x  '000098  in.) ; 
it  multiplies  by  budding  and  sporulation,  like  all  the 
other  varieties  of  this  species. 


Fig. 


The  Saccharoniyces  conglomeratiLS  of  Rees  (Fig.  1 3),  is 
rather  rare  ;  it  is  met  with  in  the  must  of  wine  towards 
the  end  of  the  fermentation.  It  has  spheroidal  cells 
of  -nfoo  millimetre  in  diameter  ('000236  in.). 


Fig.  i2.~Saccharomyces  exiguus,  X  Fig.  13.— Saccharomyces  conglomera* 
350.  tus,  X  600. 

When  the  first  cell  has  budded,  this  bud  attains  the 
size  of  the  mother-cell,  without  bejng  detached  ;  there 
originate  first  in  the  inner  angle  formed  by  two  cells, 
and  then  on  different  parts  of  their  surface,  a  consider- 
able number  of  new  cells,  which,  instead  of  forming  a 
chaplet  or  flakes,  are  entirely  conglomerated. 

The  apiculatcd  ferment  {Carpozyind)  docs  not  belong 
to  the  genus  Saccharoniyees,  according  to  the  observa- 
tions of  M.  Engcl.    This  is  the  most  abundant  alcoholic 


ALCOHOLIC  FERMENTS. 


57 


ferment.  It  is  met  with  on  all  kinds  of  fruit,  especially 
on  berries  and  stone-fruits,  as  well  as  in  the  greater 
number  of  musts  of  wines  in  process  of  fermentation. 
It  has  also  been  noticed  in  certain  kinds  of  beer,  as  that 
of  Belgium,  and  of  Obernai ;  those  of  Strasbourg  do 


Fig.  14.— Saccharomyces  apiculatus  (Rees).  Carpozyma  apic.  (Engel),  apiciilated 
ferment,  X  600. 


not  contain  it.  It  is  this  which  is  usually  the  first  to 
appear  and  bud  in  these  musts. 

The  adult  and  isolated  cells  (Fig.  14)  have  the  form 
of  an  ellipsoid,  whose  greater  diameter  is  yo%-u  of  a 
millimetre  ('000236  in.),  and  the  transverse  diameter 
one-half  of  the  larger  one.  At  each  extremity  there  is  a 
small  projection  or  minute  stalk,  which  give  the  cell  the 
appearance  of  a  lemon.  The  interior  encloses  a  spher- 
ical or  ellipsoidal  vacuole,  around  which  there  is  a  thin 
layer  of  protoplasm,  fringed  towards  the  projecting  parts. 
The  budding  cells  always  present  themselves  at  the 
extremity  of  the  projections. 

When  the  development  is  normal,  the  new  cells  extend 
in  the  direction  of  the  principal  axis  of  the  mother-cell, 
so  that  the  three  cells  form  a  longitudinal  row ;  but 
when  they  have  ceased  to  grow,  they  assume  an  ellip- 
tical form,  and  bend  back  at  the  point  of  their  insertion, 
so  that  their  longer  axis  forms  at  last  a  right  angle  with 


58 


ON  FERMENTATION. 


the  axis  of  the  mother-cell ;  one  of  the  cells  turns  to 
the  right,  and  the  other  to  the  left.  They  are  then 
detached  ;  at  this  time  they  much  resemble  the  cells 
of  Saccharomyces  ellipsoideus ;  but  we  soon  see  the  cha- 
racteristic stalks  appear. 

When  the  apiculate  ferment  is  deposited  on  damp 
plaster,  the  transformation  into  thecse  or  sporanges 
assumes  phases  very  different  from  those  which  are 
observed  in  the  various  species  of  Saccharo77iyces,  and 
resembles  the  evolutions  of  the  Protomyces  macrosporus 
studied  by  De  Bary. 

According  to  Engel,  the  apiculate  ferment  is  a  Proto- 
myces without  a  mycelium  ;  this  botanist  proposes  to 
give  it  the  name  of  Carpozyma.  Its  thecse  are  spherical, 
covered  with  a  peritheca,  and  are  hibernating.  The 
development  of  the  spores  is  very  slow,  and  the  spores 
numerous. 

Rees  met  with  a  special  form  of  ferment,  which 
accompanies  the  Saccharomyces  ellipsdideuSy  in  the  fer- 
mented musts  of  red  wine.  It  is  composed  of  elongated 
cylindrical  cells.  Although  this  ferment  has  not  been 
observed  in  the  different  evolutions  of  its  vegetative 
life,  it  has  been  thought  necessary  to  establish  for  it 
a  special  species,  under  the  name  of  Saccharofnyces 
Reesii. 

The  Saccharomyces  mycoderma  (Figs.  i6  and  17),  called 
flowers  of  wine,  or  flowers  of  beer,  ought  also,  according 
to  M.  Pasteur,  to  be  placed  among  alcoholic  ferments. 
In  fact,  although  it  does  not  act  in  this  manner,  under 
the  ordinary  circumstances  of  its  development,  when  it 
grows  on  the  surface  of  fermented  liquors,  perhaps 
because  the  alcohol  which  it  may  then  produce  is 


ALCOHOLIC  FERMENTS. 


59 


destroyed  by  a  subsequent  oxidation,  M.  Pasteur  has 
shown  that  the  Mycoderma  viiiiy  sown  in  sweetened 
water,  is  able  to  set  up  in  it  alcoholic  fermentation. 


Fig.  15. — Saccharomyces  Reesli,  ferment  Fig.  16. — Saccharomyces  mycoderma, 
of  red  wine,  x  350.  X  350. 

It  makes  its  appearance  in  all  alcoholic  liquids  ex- 
posed to  the  air,  when  the  fermentation  is  over  or  has 
become  languid.  It  grows  w^ith  great  rapidity ;  it  is 
sufficient  to  place  a  few  of  its  cells  on  the  surface  of  a 
liquid  that  easily  becomes  alcoholic,  and  we  shall  find, 
in  less  than  forty-eight  hours,  the  surface  covered  with 


Fig.  17, — Saccharomyces  mycoderma. 


a  thin  pellicle,  of  a  whitish  or  yellowish  tint,  at  first 
smooth,  and  then  wrinkled. 

M.  Engel  estimated,  by  calculation  founded  on  his  ob- 
servations, that  in  forty-eight  hours  a  cell  of  Mycoderma 
z^/;// would  produce  about  35,378  cells.  The  cells  of  this 
Mycodenna  have  various  forms — ovoid,  ellipsoidal,  and 
cylindrical,  with  rounded  extremities. 


6o 


ON  FERMENTATION. 


The  ovoid  cells  have  their  greater  diameter  about  xoVo 
of  a  millimetre,  and  their  smaller  one  i-q^-q  (about 
•000236  X  '000157  in.).  The  cylinders  have  the  longer 
diameter  about  xo~o"o^  ^i*  To'^o"  of  a  millimetre,  and  the 
lesser  one  x^Vo-  millimetre  (-00047  X  'OOOiiS  in.). 

The  cells  are  generally  poor  in  protoplasm ;  they 
show  in  their  interior  from  one  to  three  brilliant  points  of 
fatty  matter.  The  budding  is  effected  at  the  extremity, 
by  one  or  two  buds  originating  at  each  end.  Chaplets 
and  ramified  and  interlaced  flakes  are  thus  formed,  giving 
to  the  whole  the  appearance  of  a  fine  membrane. 

When  we  dilute  with  a  considerable  proportion  of 
water  the  alcoholic  liquid  on  which  the  mycoderma 


vegetates,  the  cells  undergo  great  modifications.  The 
oldest  are  weakened  and  die,  allowing  their  protoplasm 
to  escape.  The  others  lengthen,  acquiring  a  larger 
diameter  of  from  iVo  o-  to  jtoo  of  a  millimetre  ('000629 
to  '000787  in.)  ;  their  protoplasm  collects  in  different 
points  and  forms  spores.  These  are  usually  about  three 
or  four  in  number,  arranged  in  a  longitudinal  file  in  the 
cell.  The  spores  are  generally  about  yo"o  0  niillimetre 
in  diameter  ('OOOiiS  in.). 

Many  autliors  suppose  the  SaccJiai^oviyccs  mycoderma 
to  be  derived  from  the  SaccJi.  ccrcvisi<£y  one  of  the  aerial 
forms  of  which  it  represents.    This  question  docs  not 


Fig.  18. — Mucor  racemosus,  ferment  in  mass. 


4 


ALCOHOLIC  FERMENTS. 


6i 


seem  yet  definitely  settled,  although  there  is  most 
reason  to  suppose  that  it  constitutes  a  separate  and 
independent  species. 

We  shall  have  to  return  to  the  oxydizing  properties 
of  the  Mycodenna  vini  when  we  treat  of  acetic  fer- 
mentation. 

The  MtLcor  miicedo  and  the  Mitcor  racemosiis  (Fig.  i8) 
possess  the  property,  when  immersed  in  a  solution  of 
sugar,  and  protected  from  the  access  of  oxygen,  of 
transforming  or  dividing  their  mycelium  into  joints 
having  the  form  of  balls.  These  balls  are  multiplied 
by  budding,  and  excite  alcoholic  fermentation  in  sugar 
as  long  as  they  are  placed  under  these  abnormal  con- 
ditions. 

This  fact,  which  is  indisputably  proved,  gives  con- 
siderable support  to  the  theories  brought  forward  by 
some  men  of  science  as  to  the  transformation  of  fer- 
ments, from  one  to  another,  according  to  the  conditions 
under  which  they  are  placed.  We  see,  in  fact,  the 
Mucor  racemosiis  completely  change  its  mode  of  repro- 
duction when  it  is  placed,  without  access  of  oxygen,  in 
a  saccharine  medium.  Analogous  facts  are  known  to 
be  produced  in  the  case  of  other  organisms. 

These  various  kinds  of  ferments  have  been  found,  not 
only  in  the  must  derived  from  fruits,  but  also  on  the 
surface  of  their  pericarps,  to  which  they  remain  fixed  in 
a  state  of  repose,  until  by  the  concurrence  of  suitable 
circumstances,  they  are  placed  in  contact  with  the 
saccharine  liquid  contained  in  the  cells.  From  this 
moment  they  begin  to  develop  by  buds,  and  set  up,  at 
the  same  time,  alcoholic  fermentation. 

According  to  a  recent  work  by  Dr.  de  Vaureal,  the 


62 


ON  FERMENTATION. 


alcoholic  ferment,  with  its  envelope  composed  of  non- 
contractile  cellulose,  and  reproducing  by  gemmation, 
which  has  been  generally  believed  in,  is  inadmissible. 
The  supposed  budding  is  only  an  optical  delusion.  The 
utricle  of  yeast  is  allied  to  the  spermogones  of  Tulasue; 
the  granulations  or  nucleolar  elements  are  spermatia; 
these  elements  when  set  free  by  the  rupture  of  the 
utricle,  produce  new  ones. 

This  mode  of  multiplication  explains  the  facility 
with  which  the  reproductive  elements  of  yeast  can  be 
carried  by  the  air,  when  we  cannot  distinguish  in  air- 
dust  any  characteristic  globules  of  yeast. 

In  their  mode  of  multiplication,  ferments  resemble 
somewhat  the  zoospores  of  algas ;  when  they  are  not 
too  hybrid,  like  those  of  cider,  which  reproduce  a 
penicillium  and  an  aspergillus,  they  arrange  themselves 
under  the  law  of  metagenesis,  like  the  acalephae. 
Especially  in  the  genus  Hydrodiction,  w^e  notice  a  great 
similarity. 

In  fact,  we  see  in  this  zoospores  of  two  sorts;  the 
greater  ones  (macrogonidia)  are  true  spores  ;  they  have 
a  rapid  development  and  direct  evolution  ;  the  smaller 
ones  (microgonidia)  have  a  slow  development  ;  they  do 
not  reproduce  the  plant,  but  produce  in  their  interior 
true  zoospores.    These  are  young  spores,  like  ferments. 


FERMENTS. 


63 


ACTUAL  COMPOSITION  OF  FERMENTS. 

Before  commencing  this  subject,  in  which  we  shall 
have  to  consider,  among  other  questions,  the  chemical 
modifications  which  take  place  in  ferments  under  the 
different  conditions  in  which  they  may  be  placed,  we 
ought  to  give  a  summary  of  the  results  obtained  by 
experimentalists  who  have  devoted  themselves  to  the 
study  of  the  chemical  composition  of  these  organisms. 

Much  has  been  done  in  this  respect.  Thus  Schloss- 
berger  has  published  very  careful  researches  on  the 
actual  elementary  composition  of  the  two  kinds  of  fer- 
ment of  beer,  freed  as  far  as  possible,  by  washing  and 
decanting,  from  the  impurities  which  are  found  in  the 
crude  yeast. 

This  observer  found  as  the  mean  of  two  analyses  : — 


SURFACE 
YEAST. 


SEDIMENTARY 
YEAST. 


.  Results  ascertained, 
the  ashes  having  ^ 
been  removed 


Carbon  .  49*9 
Hydrogen  .  6  6 
)  Nitrogen  .  I2*i 
Oxygen  .31*4 
Ashes        .  2*5 


.11-6. 


.  48-0 
.  6-5 
.  9-8 
.  357 
.  3*5 


Messrs.  Mitschcrlich,  Mulder,  and  Wagner  have  pub- 
lished independently  the  following  results  : — 


64 


ON  FERMENTATION. 


SURF.  SURF. 

YEAST.  YEAST. 

(Mulder.)  (Wagner.) 

50-8  49*8 

7*2  6-8 

II'l  9*2 


M.  Dumas  (Traite  de  Chimie),  finds : — 

.  50-6 

.  7-3 
.  15-0 

•  27*1 

This  result  differs  from  the  others  by  a  larger  pro- 
portion of  nitrogen  ;  but  we  can  understand  variations 
in  the  composition  of  such  a  product  as  yeast,  which  is 
constantly  undergoing  a  process  of  chemical  evolution. 
Thus,  the  smaller  percentage  of  nitrogen  and  carbon 
furnished  by  analyses  of  the  sedimentary,  as  compared 
with  that  of  the  surface  yeast,  is  easily  explained,  if  wc 
keep  in  mind  the  fact  that  the  former  remains  much 
longer  in  contact  with  the  liquid  after  the  fermentation. 
Phenomena  dependent  on  spontaneous  change  of  con- 
dition may  then  take  place,  transforming  into  soluble 
principles  a  portion  of  the  nitrogenous  albuminoid 
products  of  the  protoplasm,  and  allowing  them  to 
escape  into  the  surrounding  liquid. 

Schlossberger  also  endeavoured  to  isolate  the  various 
ultimate  principles  contained  in  yeast.  By  treating  it 
witli  a  very  weak  solution  of  potash,  filtering  and  neu- 


The  ashes  not 
separated. 


SED. 

YEAST. 

(Wagner.) 

f  Carbon  44*4 

Hydrogen  6*o 

Nitrogen  9*2 
Sulphur 

Oxygen  35-8 


SURF. 
YEAST. 

(Mitsch.) 

47-0 

6-6 

lO'O 

0-6 


Carbon  . 
Hydrogen 
'Nitrogen 
Oxygen 
Sulphur 
Phosphorus 


I  .  .  . 


ACTUAL  COMPOSITION  OF  FERMENTS. 


65 


tralizing  the  liquid  by  an  acid,  he  obtained  a  floccose 
white  precipitate,  free  from  ashes,  which,  when  analyzed 
for  its  elements,  gave  the  following  numbers : — 

MEAN  OF  TWO  ANALYSES. 


Carbon  55*5 

Hydrogen  7*5 

Nitrogen  .  .  ,  ,  •  I3*9 
Sulphur  o* 


The  nitrogen  is  here  present  in  too  small  a  propor- 
tion for  a  normal  albuminoid  compound.  The  analysis 
agrees  tolerably  well  with  that  made  by  me  of  hemi- 
protein,  one  of  the  products  of  the  decomposition  of 
albumin,  by  dilute  sulphuric  acid  {see  the  chapter  on 
albuminoid  substances).  This  similarity  is  the  more 
striking,  since  the  hemi-protein  is  also  soluble  in  dilute 
alkalis,  and  precipitated  by  acids.  The  precipitate, 
when  well  washed,  yields  no  ashes  after  combustion. 
On  the  other  hand,  it  is  very  probable  that  the  yeast 
acts  on  albuminoid  substances  by  splitting  them  up 
progressively,  of  which  we  shall  find  proofs  farther  on. 

By  saturating  the  yeast  with  acetic  acid  and  pre- 
cipitating the  filtered  liquor  by  ammonium  carbonate, 
Mulder  obtained  a  principle  nearer  in  its  composition 
to  albumin,  and  which  gives, — 

Carbon  53-3 

Hydrogen  .  ,  .  ,  .7*0 
Nitrogen  i6*o 

We  may  therefore  admit  the  presence  of  one  or  more 
albuminoid  substances  in  the  yeast-cell ;  in  this  respect 
it  does  not  difi*er  from  other  vegetable  cells. 

The  residuum  insoluble  in  potass,  in  Schlossberger's 


66 


ON  FERMENTATION. 


experiment,  was  then  saturated  with  acetic  acid  and 
water.  It  then  shows  that  it  has  a  composition  alHed 
to  that  of  cellulose  : — 

Carbon  ......  44*9 

Hydrogen  67 

Nitrogen  0*5 

Remaining  ashes  .       .       ,       .  I'l 

This  cellulose,  boiled  with  sulphuric  acid,  is  easily 
converted  into  fermentable  sugar.  According  to  Liebig, 
it  is  not  soluble  in  ammoniacal  cupric  oxide.  It 
seems,  therefore,  to  differ  from  normal  cellulose,  soluble 
in  ammoniacal  cupric  oxide,  and  which  dilute  acids  do 
not  transform  into  sugar. 

Payen  (Memoire  des  Savants  Etrangers,  vol.  9,  p.  32) 
gives  the  following  direct  analysis  for  yeast : — 

Nitrogenous  matter  .  .  •  6273 
Cellulose  (envelopes)  .  .  .  22*37 
Fatty  matter  .  •  •  .  2'io 
Mineral    „         ....  5'8o 

Other  experimentalists,  such  as  Pasteur  and  Liebig, 
employing  the  methods  of  separation  usual  in  the 
analysis  of  the  higher  plants,  have  found  only  18*5 
per  cent,  of  pure  cellulose  in  fresh  yeast.  If  we  admit 
that  the  nitrogen  (i  r8  to  12*5  per  cent.)  contained  in 
the  yeast,  forms  an  integral  part  of  the  albuminoid 
matter,  we  can  caculate,  by  simple  rule  of  three,  that 
the  yeast  contains  about  60  per  cent,  of  proteids  and 
nearly  40  per  cent,  of  hydrocarbons. 

This  method  of  calculating  the  actual  composition  of 
yeast  is  not  quite  legitimate.  We  find,  in  fact,  in  the 
washings  of  fresh  yeast,  performed  in  ice-cold  water, 
noticeable  quantities  of  tyrosine,  leucine,  &c.,  which 

J 


ACTUAL  COMPOSITION  OF  FERMENTS.  6/ 

contain  less  nitrogen  than  albuminoid  matters  (lo  and 
77  per  cent.,  instead  of  from  15*5  to  16). 

As  direct  analyses  give  only  1 8*5,  or  at  most  30,  per 
cent,  of  cellulose,  one  is  led  to  suppose  that  in  the 
yeast-cell  other  hydrocarbons  are  found,  more  easily 
attacked  by  acids  and  alkalis  than  is  true  cellulose. 
This  opinion  is  corroborated  by  the  production  of 
alcohol  during  the  digestion  of  yeast,  without  the 
addition  of  sugar,  although  one  cannot  discover  in  it 
the  presence  of  glucose.  On  the  other  hand,  we  find  in 
the  extract  of  spontaneously  decomposed  yeast  notice- 
able quantities  of  a  special  gummy  substance  (Bechamp, 
Schutzenberger).  The  origin  of  this  gum,  if  it  does 
not  pre-exist  fully  formed  in  the  fresh  cell,  can  only 
be  attributed  to  the  splitting  up  of  a  compound  of 
the  family  of  glucosides,  or  to  a  molecular  transfor- 
mation of  an  insoluble  hydrocarbon  substance,  different 
from  cellulose.  Pasteur  (Compt.  Rend.,  vol.  48,  p.  640) 
obtained  20  per  cent,  of  sugar  by  boiling  yeast  with 
dilute  sulphuric  acid. 

We  owe  to  Mitscherlich  (Ann.  der  Chemie  und  Phar 
macie,  vol.  56  )  some  excellent  analyses  of  the  ashes  of 
yeast.  The  dried  matter,  placed  in  a  silver  crucible, 
itself  placed  in  one  platinum,  was  burnt  in  a  glass  tube, 
in  a  current  of  oxygen ;  the  distillation  of  the  organic 
matter  was  commenced  in  an  atmosphere  of  carbon 
dioxide,  and  the  combustion  was  finished  in  oxygen. 

Quantity  of  ash  of  the  yeast : — 

SURF.  YEAST.  SED.  YEAST.  SURF.  YEAST. 

(Wagner  and  (Schlossberger.)  (Bull.) 

Schlossberger.) 

2*5  per  cent.      3*5  to  4  per  cent.       8*9  per  cent. 


SED.  YEAST. 

(Mitsch.) 
77  per  cent. 


68 


ON  FERMENTATION, 


SED.  YEAST.  SED.  YEAST. 

(Mitsch.)  (Wagner.) 
7 '5  per  cent.  5*3  per  cent. 

Taking  only  the  results  obtained  by  Mitscherlich, 
which  appear  the  most  reliable,  there  would  be  no 
difference  with  respect  to  the  quantities  of  mineral 
matters  between  the  two  kinds  of  yeast.  The  small 
proportion  of  mineral  matter  found  by  Schlossberger 
may  have  resulted  from  the  fact  that  this  experimen- 
talist washed  his  yeast,  an  operation  which,  as  Bechamp 
has  shown,  gives  rise  to  a  continuous  elimination  of 
phosphates. 

The  ash  of  the  ferment  gives  the  following  per- 
centage : — 


SURF.  PERM. 

SED.  PERM. 

SURF.  FERM. 

(Mitscherlich.) 

OF  PALE 

ALE  (Bull.) 

Phosphoric  acid 

.  53*9 

.  59*4 

•  547 

Potass 

.      .  39-8  . 

.  28-3 

.  35*2 

Soda       .  , 

Magnesia. 

.  6-0 

.  8-1 

.  4-1 

Lime       ,  . 

.  I'O 

.  4*3 

.  4*5 

Sihca 

.  traces 

Iron  Oxide 

.  0-6 

Sulphuric  acid  . 

Hydrochloric  acid 

.  O'l 

The  predominant  elements  are,  therefore,  phosphoric 
acid  and  potass,  together  with  a  little  magnesia  and 
lime. 

The  analyses  of  Mitscherlich  may  be  thus  calcu- 
lated :— 


ACTUAL  COMPOSITION  OF  FERMENTS.  69 

SURF.  YEAST.        SED.  YEAST, 

Phosphoric  acid  .      •      .      .41*8  ,       .  39*5 

Potassa  39*8  .       .  28*3 

Soda  —  •       .  — 

Magnesium  phosphate,  )  ^^.g  ^^.^ 

(Mg.3  2P04)  .       .   )   •       •  •  • 
Calcium  phosphate  \ 

(Ca3  2  PO4)    .       .  >  * 


2-3      .      .  97 


The  estimates  which  we  have  just  given  of  the  direct 
and  elementary  composition  of  the  yeast  are  still,  as 
we  see,  incomplete ;  and  this  question  is  worth  atten- 
tively studying  again. 

This  opinion  has  already  been  brought  forward  by 
Pasteur,  who  thus  expresses  himself  (Ann.  Chim.  Phys. 

(3)  58,  p.  403):— 

"  Yeast  contains  several  nitrogenous  substances,  and 
also  some  not  nitrogenous — substances  distinct  from  each 
other.  It  would  be  interesting  to  study  this  subject. 
I  have  found  that  we  should  arrive  at  useful  results  by 
examining  separately  the  action  of  water,  of  dilute 
sulphuric  acid,  and  of  potass.  I  think  that  an  examina- 
tion of  yeast,  made  for  this  purpose,  and  of  the  different 
materials  which  compose  it,  might  reveal  the  secrets  oi 
certain  changes  which  are  observed  in  the  nature  of  the 
extract  of  the  fermented  liquid." 

Notwithstanding  the  insufficiency  of  the  data  of  this 
question,  we  see  that,  considered  qualitatively,  the  cells 
of  yeast  resemble  other  cells  which  enter  into  the  com- 
position of  larger  plants,  and  of  fungi  in  general ;  they 
have  envelopes  formed  of  cellulose  in  different  degrees 
of  evolution ;  and  their  contents  are  composed  princi- 
pally of  an  albuminous  protoplasm,  of  hydrocarbons 


70 


ON  FERMENTATION. 


analogous  to  gum,  with  fatty  and  even  resinous  sub- 
stances. Indeed,  yeast  contains  a  small  quantity  of 
bitter  resin  soluble  in  alkalis. 

When  examined  quantitatively,  it  presents  a  special 
character,  and  is  very  rich  in  nitrogen,  far  more  so  than 
the  vegetable  tissues  in  general.  Thus  in  fungi,  Messrs. 
Schlossberger  and  Doepping  found  in  1 00  grammes  of 
dry  matter  the  following  quantities  of  nitrogen  : — 

CANTHARELLUS  KUSSULA.  LACTAKIUS 

(CHANTARELLE).  DELICIOSUS. 

gram.  gram.  gram. 

Nitrogen,  per  cent.  .  3*22       .       .       .4*25      .       .  4*68 


BOLETUS  EDULIS.  AGARICUS  CAMPESTKIS. 

gram.  gram. 
Nitrogen,  per  cent.     .  47     .       .       .       .  7*26 

The  composition  of  various  fungi,  according  to  Payen, 
is  the  following: — 

CULTIVATED     MORELLS.        WHITE  BLACK 
MUSHROOMS.  TRUFFLES.  TRUFFLES. 

Water    .       .       .       .  qi'oi    .    90        .  72*34        .  72 

Nitrogenous  com- 
pounds, with  a 

trace  of  sulphur  .  4'68  ,  4*4  .  g'gG  .  876 
Fatty  matter  .  .  0*40  ,  0*56  .  0*44  .  0*56 
Cellulose,  dextrin, 

sugars,  tertiary 

matter       .       .       .    5-45     .    3-68     .  I5'i6i       .  7-59 

Salts,  Alkalne,  cal- 
cic, magnesic, 
silicic  phosphates 

and  chlorides     .       .    o'^G     .    1*36     .    2*  10        .  2*07 
Nitrogen  per  cent. 

of  the  dry  matter       .    7*33     .    —       .    1*532       .  1*35 


ACTUAL  COMPOSITION  OF  FERMENTS.  71 

Cultivated  mushrooms  are,  therefore,  almost  as  rich  in 
albuminoid  matter  as  yeast,  since  100  parts  of  dry 
matter  contain  52  parts  of  these  substances.  Yeast 
contains  60. 


72 


ON  FERMENTATION. 


CHAPTER  V. 

FUNCTIONS  OF  YEAST. 

Yeast  is  a  living  organism  belonging  to  the  family  of 
fungi,  genus  SaccharomyceSy  destitute  of  mycelium, 
capable  of  reproduction,  like  all  the  elementary  fungi, 
by  buds  and  spores  ;  its  composition,  as  we  have  just 
seen,  singularly  resembles  that  of  other  vegetable  tissues, 
and  especially  of  the  plants  of  the  same  family.  The 
examination  of  its  biological  functions,  studied  more 
particularly  in  their  chemical  aspect,  shows  us  clearly 
that  this  elementary  form  of  life  does  not  differ  in 
essentials  from  other  elementary  cells,  unprovided  with 
chlorophyll,' whether  isolated  or  in  groups,  and  belong- 
ing to  the  more  complex  organs.  It  breathes,  trans- 
forms and  modifies  its  proximate  principles  in  a 
continuous  manner,  and  certainly  in  the  same  way  as 
other  cells  ;  like  these,  it  can  be  multiplied  by  buds  and 
spores.  The  only  important  and  decidedly  distinctive 
character  which  seems  to  render  it  a  form  of  life 
absolutely  apart  from  other  forms  in  creation,  was 
removed  from  it  by  M.  Lechartier  and  M.  Bellamy, 
when  these  chemists  succeeded  in  establishing  that  the 
cells  of  fruits,  seeds,  and  leaves,  and  even  animal  cells, 
are  capable  of  changing  sugar  into  alcohol  and  carbon 
dioxide. 

From  this  time,  therefore,  the  accurate  study  of  the 


FUNCTIONS  OF  YEAST. 


73 


biological  functions  of  yeast  no  longer  appears  to  us 
as  an  isolated  chapter  in  the  midst  of  those  which  com- 
pose general  physiology,  but,  on  the  contrary,  as  a  par- 
ticular instance,  from  which  we  may  draw  important 
conclusions  as  to  the  chemical  phenomena  of  living 
organisms  in  general.  Yeast  offers  this  immense  advan- 
tage to  the  observer,  that  it  allows  him  to  make  all 
kinds  of  experiments  on  it  with  great  facility.  It  is  like 
clay,  which  can  be  moulded  at  will,  composed  of  one 
and  the  same  kind  of  elementary  cells,  which  enables 
us  to  avoid  the  complications  due  to  the  intervention  of 
complicated  phenomena.. 

Normal  Conditions  of  the  Life  of  Yeast. — The  condition 
which  we  shall  call  normal  in  the  life-history  of  yeast 
are  those  in  which  this  form  of  life  develops  itself,  and 
increases  with  the  greatest  activity  and  energy.  They 
are  of  two  orders,  physical  and  chemical. 

With  respect  to  physical  conditions,  we  have  only  to 
notice  the  temperature.  The  temperature  most  favour- 
able to  the  nutrition  of  yeast  is  also  that  which  is  found 
advantageous  to  other  cellular  vegetable  organs ;  between 
25°  C.  and  35°  C.  (77"  and  95°  R).  Above  and  below 
these  limits,  the  vital  manifestations  do  not  cease  until 
we  descend  below  9°  C.  (49'6''  F.)  or  rise  above  60°  C. 
(140°  R),  the  temperature  at  which  albuminoid  principles 
begin  to  coagulate. 

As  to  the  chemical  conditions,  and  the  most  favour- 
able composition  of  the  medium  in  which  this  organism 
is  to  live  and  multiply,  they  are  not  so  simple. 

We  owe  to  the  learned  and  patient  labours  of  Pasteur  on 
this  question  the  best  information  which  we  possess  on  this 
part  of  the  subject.  He  has  been  followed  in  this  interest- 


74 


ON  FERMENTATION. 


ing  investigation  by  some  of  his  pupils  (Duclaux,  Raulin), 
of  whose  researches  we  shall  have  to  speak  further  on. 

We  can  see,  a  priori^  that  the  most  favourable  medium 
is  that  which  contains  the  most  appropriate  nutritive 
elements.  These  elements  ought  to  be  those  which  we 
find  in  the  organism  of  the  cell,  or,  at  least,  principles 
susceptible  of  allowing  the  cell  to  form  with  them  by 
synthesis,  its  immediate  component  parts.  Thus  we 
have  seen  that  yeast  contains  water  in  greater  or  less 
proportion,  mineral  salts,  especially  potassium,  magne- 
sium, and  calcium  phosphates.  Water  and  the  alkaline 
and  alkaline-earthy  phosphates  will  therefore  necessarily 
form  an  integral  part  of  the  nutritive  medium.  We 
find,  besides,  a  great  proportion  of  nitrogenous  sub- 
stances, either  albuniinous  or  otherwise.  The  food  of 
yeast  ought,  therefore,  to  include  nitrogen.  A  question 
then  presents  itself :  Under  what  form  ought  nitrogen 
to  be  supplied  to  the  cell  in  order  to  be  assimilated  t 
Experiment  alone  can  reply. 

We  know,  by  the  elegant  researches  of  M.  Boussin- 
gault,  that  the  higher  plants  are  not  able  to  absorb  the 
free  nitrogen  of  the  atmosphere.  This  observer  caused 
a  certain  known  weight  of  seeds,  the  average  amount  of 
nitrogen  in  which  he  had  previously  ascertained,  to 
germinate  in  a  calcined  silicious  soil,  and  watered  them 
with  pure  water.  When  the  plant,  in  spite  of  these 
unfavourable  conditions,  had  attained  a  certain  develop- 
ment, he  ascertained  the  total  quantity  of  nitrogen  con- 
tained in  the  whole.  The  weight  of  this  did  not  exceed 
the  original  quantity  contained  in  the  seed.  Yeast  forms 
no  exception  to  this  rulc; 

Assimilation  of  the  Nitrogen  of  Nitrates, — Agricultural 


FUNCTIONS  OF  YEAST. 


75 


experiments  made  on  a  large  scale,  and  repeated  for 
successive  years,  as  well  as  those  made  in  chemical 
laboratories,  have  established  the  efficacy  of  nitrates, 
and  salts  of  ammonium,  among  the  nitrogenous  aliments 
utilized  by  plants  in  general. 

The  importance  of  saltpetre  as  a  manure,  has  been 
long  known  ;  however,  with  respect  to  biological  phe- 
nomena, it  is  a  question  whether  the  nitrate  introduced 
into  the  soil  does  not,  before  it  reaches  the  plant  which 
makes  use  of  nitrogen,  undergo  changes  which  reduce 
it  to  another  form,  that,  for  instance,  of  ammoniacal  salts. 

M.  Boussingault  has  given  a  complete  solution  to 
this  question  by  experimenting  on  plants  sown  in  a 
medium  or  a  soil  absolutely  deprived  of  organic  matter, 
and  in  which  the  reduction  of  nitrates  is  impossible. 
The  following  table  gives  an  idea  of  the  influence  of 
nitrates  on  ves^etation  in  a  barren  soil: — 


Weight  of  the 
dry  produce, 

the  seed 

being  i. 


1.  When  the  soil 
had  received  no- 
thing ...  3-6 

2.  When  the  soil 
had  received 
phosphates, 
ashes,  and  potas- 
sium nitrate       .  198*3 

3.  When  the  soil 
had  received 
p  hosphates, 
ashes,  and  potas- 
sium bicarbonate.  4*6 


Vegetable 

matter 
elaborated. 


0*285 


21*111 


0*391 


Carbon  dioxide 
decomposed  in 
24  hours. 


2*45 


182*00 


Acquired  by  the  plants 
during  86  days 
of  vegetation. 

Carbon.  Nitrogen. 


grammes. 


3*42 


0-I44 


8-446 


0*156 


0*0023 


o*i666 


0*0027 


76 


ON  FERMENTATION. 


These  numbers  show  such  important  differences  be- 
tween the  results  obtained  in  the  same  time,  with  or 
without  the  use  of  nitrates,  all  things  besides  being 
equal,  that  an  error  of  interpretation  is  impossible. 

We  may  then  say :  In  the  larger  plants  the  soluble 
nitrate  penetrates  into  the  organism  in  a  state  of  solu- 
tion, and  there  it  undergoes  changes  which  finally 
bring  out  its  nitrogen,  unSer  the  form  of  albuminoid 
matter. 

Is  it  the  same  with  yeast  ?  Can  it  elaborate  its  pro- 
teic  matter  by  decomposing  nitrates  ? 

This  question  has  been  discussed.  On  one  side, 
Dubrunfant  (Comp.  Rend,  de  TAcad.,  vol.  73,  pp.  200, 
263)  says  that  he  has  observed  a  greater  activity  as  a 
ferment  after  the  addition  of  potassium  nitrate. 

On  the  other  side.  Ad.  Mayer  (Lehrbuch  der 
Gahrungs-Chemie,  1874)  affirms  that  he  has  obtained 
only  negative  results  in  a  whole  course  of  researches, 
directed  to  this  end.  Schaer  arrived  at  the  same  results 
as  Mayer. 

This  decided  difference  between  the  phenomena  of 
the  nutrition  of  yeast  and  of  larger  plants  is  still 
more  remarkable,  since  simple  organisms,  much  allied 
to  the  Saccharomyces,  act  upon  nitrates  exactly  in  the 
same  way  as  terrestial  plants  of  a  higher  order. 

Thus,  the  mildews  which  vegetate  on  the  surface  of 
liquids  derive  considerable  nourishment  from  the  dis- 
solved nitrate. 

Admitting  the  observations  of  Mayer  to  be  correct, 
we  are  compelled  to  interpret  these  negative  results  in 
the  following  manner  : — 

In  order  to  assimilate  the  nitrogen  of  a  nitrate,  the 


FUNCTIONS  OF  YEAST. 


77 


plant  must  first  reduce  it.  Therefore,  either  the  cell  of 
the  Saccharomyces  does  not  possess  this  reducing  power 
over  the  nitrates  which  we  find  in  other  isolated  cells, 
forming  an  integral  part  of  more  complex  organisms, 
or  else  the  experiments  have  been  made  under  con- 
ditions in  which  this  reducing  power  has  not  been  able 
to  manifest  itself.  However  this  may  be,  we  have  not 
arrived  at  a  final  decision  respecting  the  assimilation 
of  the  nitrogen  of  nitrates  ;  and  before  we  determine 
definitely  in  the  negative  it  will  be  advisable  to  vary  the 
experiments. 

Assimilation  of  the  Nitrogen  of  Ammoniacal  Salts. — 
Agriculturists  generally  agree  in  recognizing  the  efficacy 
of  ammoniacal  salts  in  vegetation.  The  experiments 
of  Sir  H.  Davy,  Kuhlmann,  J.  Pierre,  Lawes,  and 
Gilbert  lead  us  to.  the  same  opinion.  It  appears  proba- 
ble, according  to  all  the  facts,  that  ammoniacal  salts 
may  concur  in  the  nutrition  of  plants,  although  it  is 
recognized  that,  with  an  equal  weight  of  nitrogen,  they 
act  in  a  less  favourable  manner  than  the  nitrates.  The 
experiments  of  M.  Bouchardat  (Memoire  sur  ITnfluence 
des  Composes  Ammoniacaux  sur  la  Vegetation),  those  of 
M.  Cloez  (Legons  de  la  Soc.  Chim.,  1861,  p.  167)  tend, 
on  the  contrary,  to  establish,  1st,  that  the  solutions  of 
ammoniacal  salts,  usually  employed,  do  not  supply  to 
vegetables  the  nitrogen  which  they  assimilate ;  2ndly, 
that  if  solutions  at  the  rate  of  toVo>  ^^en  of  xo^ 0 o» 
these  salts  are  absorbed  by  roots,  they  act  as  energetic 
poisons,  and  kill  the  plant  rapidly.  M.  Cloez  cites,  on 
this  subject,  many  instances,  which  leave  no  doubt  as  to 
the  reality  of  the  fact. 

If  this  be  the  case,  we  must,  in  order  to  make  these 


78 


ON  FERMENTATION. 


apparently  discordant  results  agree,  either  admit  that 
the  ammonia,  in  order  to  be  absorbed  usefully,  and  with- 
out danger  to  the  plant,  ought  to  be  presented  to  it 
under  a  special  form,  perhaps  very  diluted,  or  in  a 
peculiar  state  of  combination  ;  or  else  it  must  previ- 
ously undergo,  in  the  soil  itself,  a  transformation  into  a 
nitrate.  This  interpretation,  due  to  M.  Cloez,  and 
which  is  the  reverse  of  the  ideas  of  M.  Kuhlmann  (for 
this  observer  maintains  that,  on  the  contrary,  the 
nitrates  employed  as  manures  are  only  active  if  they 
are  transformed,  in  the  soil,  into  ammoniacal  salts),  is 
by  no  means  contradictory  to  what  w^e  know  of  the 
phenomena  of  nitrification. 

Let  us  return  from  the  higher  forms  of  plants  to 
ferments,  as  we  have  already  done  before.  M.  Pasteur 
was  the  first  to  study  the  influence  of  ammoniacal  salts 
on  the  development  and  nutrition  of  the  Saccharo- 
myces. 

After  having  ascertained,  by  experiments  made  on  a 
large  scale  in  fermentation,  carried  on  commercially,  that 
the  ammonia  of  the  ammoniacal  salts  contained  in  the 
juices  employed  disappeared  during  the  fermentation, 
without  disengaging  any  sensible  quantity  of  nitrogen, 
he  adopted  the  following  experiment,  which  may  serve 
as  a  type  for  all  the  rest. 

In  a  solution  of  pure  sugar-candy  (we  shall  presently 
see  that  sugar  and  its  analogues  are  the  food  necessary  for 
yeast)  we  place,  first,  an  ammoniacal  salt — for  instance, 
some  ammonium  tartrate — and  then  the  mineral  matter 
which  enters  into  the  composition  of  yeast,  and  add  to 
this  a  slight,  we  may  say  imponderable,  quantity  of 
globules  of  fresh  yeast.    The  globules  sown  under  these 


FUNCTIONS  OF  YEAST. 


79 


conditions  are  developed,  multiply,  and  excite  fermenta- 
tion of  the  sugar,  while  the  mineral  matter  dissolves  by 
slow  degrees,  and  the  ammonia  disappears.  In  other 
words,  the  ammonia  is  transformed  into  the  complex 
albuminoid  matter  which  enters  into  the  composition  of 
the  yeast,  whilst  the  phosphates  supply  to  the  new 
globules  their  mineral  principles  :  as  for  the  carbon,  it 
is  evidently  furnished  by  the  sugar. 

This,  for  example,  is  the  composition  of  one  of  the 
liquids  employed, — 

lo  grammes  of  pure  sugar-candy. 

Ashes  of  one  gramme  of  yeast,  obtained  by  means 
of  a  cupel-furnace, 

o*i  gramme  of  ammonium  dextro-tartrate, 

Traces  of  washed  beer-yeast,  of  about  the  size  of  a 
pin's  head,  in  a  fresh  condition,  damp,  losing  80  per  cent, 
of  water  at  100°  C.  (212°  F.) 

In  such  a  mixture,  the  vessel  being  filled  up  to  the 
neck,  and  well  stopped,  or  furnished  with  a  gas  tube 
dipping  into  pure  water,  the  fermentation  began.  After 
from  twenty-four  to  thirty-six  hours  the  liquor  began  to 
give  evident  signs  of  fermentation,  by  a  disengagement 
of  microscopic  bubbles,  which  announced  that  the  liquid 
was  already  saturated  with  carbon  dioxide.  On  the 
following  days  the  troubling  of  the  liquor  increased 
progressively,  as  well  as  the  disengagement  of  gas, 
which  was  considerable  enough  for  the  froth  to  fill  the 
neck  of  the  flask.  A  deposit  began,  by  degrees,  to 
form  at  the  bottom  of  the  vessel.  A  drop  of  this 
deposit,  examined  under  the  microscope,  shows  a  beau- 
tiful example  of  yeast,  very  much  ramified,  extremely 
young  in  appearance  ;  that  is  to  say,  the  globules  are 


80 


ON  FERMENTATION. 


swollen,  translucent,  not  granulated,  and  we  distinguish 
among  them,  with  surprising  facility,  each  globule  of 
the  small  quantity  of  yeast  sown  at  the  commencement 
of  the  experiment.  These  latter  globules  have  a  thick 
envelope,  defined  by  a  darker  circle  ;  their  contents  are 
yellowish ;  they  are  granular  ;  but  the  manner  in  which 
they  are  sometimes  surrounded  by  the  newly  formed 
globules  shows  very  clearly  that  they  have  given  rise  to 
those  among  the  latter  which  form  the  first  links  of  the 
chaplets.  If  the  observations  are  made  in  the  earlier 
days  of  the  formation,  during  the  evening,  and  by  gas- 
light, the  old  globules  are  distinguishable  among  the 
infinitely  more  numerous  young  ones,  as  you  would  dis- 
tinguish a  black  ball  in  the  midst  of  a  great  number  of 
white  ones. 

This  fundamental  experiment  proves,  then,  that  yeast 
can  bud,  multiply,  live  and  feed  in  a  medium  in  which 
nitrogen  is  only  represented  by  ammoniacal  salts. 

Further,  M.  Pasteur  draws  this  conclusion  from  it 
that  yeast  causes  with  ammonia  the  synthesis  of  albu- 
minoid substances.  In  order  to  test  this  conclusion, 
M.  Pasteur  weighed  the  ammonia  which  remained  in  the 
liquid  some  weeks  afterwards,  and  found  it  less — slightly 
so,  it  is  true  ('0062  grammes).  The  weight  of  the  newly 
formed  yeast  rose  to  '043  grammes.* 

In  his  last  memoir  on  the  origin  of  muscular  force, 
(Ann.  Chim.  Phys.  [4],  vol.  23,  p.  5,  1871),  Licbig  ener- 
getically attacks  the  conclusions  and  results  announced 
by  Pasteur.  lie  absolutely  denies  the  formation  of 
yeast,  and  its  increase  in  weight,  under  the  conditions 
of  Pasteur's  experiments,  saying  that  he  had  never 
succeeded  in  realizing  it ;  while,  by  substituting  for  the 


FUNCTIONS  OF  YEAST. 


8i 


ammoniacal  salts  water  in  which  yeast  had  been  washed, 
the  formation  of  fresh  yeast  was  very  evident.  One  of 
the  arguments  of  Liebig,  on  which  he  rehes  most,  is  the 
absence  of  sulphur  in  the  nutritive  medium  used  by 
Pasteur ;  the  albuminous  substances  contain  some,  and, 
therefore,  yeast  cannot  elaborate  proteinic  matter  under 
these  conditions. 

Let  us  remark,  on  this  head,  that  sulphur  occupies 
but  little  space  in  the  complex  molecule  of  albumin, 
and  that  nothing  proves  that  this  body  is  indispensable 
to  the  constitution  of  albumin. 

In  reply  to  this  attack,  M.  Pasteur  could  only  repeat, 
with  forcible  conviction,  his  former  affirmations,  and 
propose  to  his  opponent  to  have  the  facts  investigated 
by  scientific  referees.  We  will  dwell  no  longer  on  this 
dispute,  which  was  unfortunately  terminated  by  the 
death  of  one  of  the  most  distinguished  chemists  of  our 
age.  We  will  only  remark  that  fresh  facts,  studied  with 
great  attention,  especially  by  M.  Raulin,  have,  by 
analogy,  given  complete  comfirmation  to  M.  Pasteur  s 
opinions  concerning  the  nutrition  of  simple  organisms 
in  general,  and  yeast  in  particular.  We  shall  allude  to 
these  facts  later  on. 

M.  Pasteur's  experiment,  as  described  by  him  in  his 
memoir  on  alcoholic  fermentation  (Ann.  de  Chimie  et 
de  Phys.,  vol.  58  [3],  p.  390),  may  be  liable  to  this 
criticism — that,  in  proportion  to  the  small  quantities  of 
yeast  at  first  employed,  the  weight  of  ammonia  that 
had  disappeared,  and  that  of  the  newly  formed  yeast, 
are  very  minute  ; '  and  that  they  might  be  considered 
as  falling  within  the  limits  of  experimental  errors,  if  one 
did  not  know  the  great  ability  of  this  eminent  observer. 


82 


ON  FERMENTATION. 


These  very  small  initial  quantities  of  yeast  were 
employed  to  avoid  the  suspicion  that  the  nourishment 
f  the  new  cells  had  been  effected  at  the  expense  of  the 
soluble  principles  excreted  from  the  old  ones  (the  water 
in  which  yeast  has  been  washed  is,  in  fact,  very  effectual 
in  giving  activity  to  the  multiplication  of  the  cells  of 
Saccharomyces),  The  experiments  made  by  Duclaux 
in  a  different  manner  prove  that  these  objections  have 
no  real  foundation,  and  that  the  ammonia  of  the 
medium  actually  disappears. 

This  skilful  chemist  introduced  into  a  certain  volume 
of  a  solution  of  sugar,  2*501  grammes  of  yeast,  containing 
0*215  grammes  of  nitrogen;  the  liquid  also  contained 
I  gramme  of  ammonium  tartrate,  corresponding  to  0*152 
grammes  of  ammonia.  After  the  fermentation,  2*326 
grammes  of  yeast,  containing  0*148  grammes  of  nitrogen, 
were  obtained  from  it.  The  liquid  contained  0*055 
grammes  of  ammonia,  and  0*170  grammes  of  nitrogen 
in  the  form  of  organic  compounds;  which  gives  the 
following  balance  of  nitrogen  : — 


Nit.  before  Nit.  after 

fermentation.  fermentation. 

In  the  yeast  0*215  •      •  .0*148 

In  the  composition  of  ammonia  .       .0*152  .       ,      •  0*045 

Under  the  form  of  nitrogenous 
organic  matter,  dissolved  in 

ihe  liquid   ...  0*120 

Total  •      •      •      .0-367  0*363 


The  two  amounts  agree,  within  an  error  of  4 
milligrammes  ('06  grains  Eng.).    We  see  clearly  that 


FUNCTIONS  OF  YEAST. 


83 


three-fourths  of  the  ammonia  has  disappeared,  and 
we  find  it  in  the  yeast  and  the  surrounding  Hquid, 
under  the  form  of  nitrogenous  organic  combinations. 
This  experiment  having  since  received  the  confirmation 
of  every  well-observed  fact,  we  may  admit,  with  confi- 
dence, that  yeast  can  effect  the  synthesis  of  its  proteinic 
materials,  at  the  expense  of  sugar  and  ammonia. 

M.  Mayer  has  proved,  as  a  complement  of  the 
experiments  of  M.  Pasteur  and  M.  Duclaux,  that  the 
ammonium  tartrate  may  be  replaced  by  other  ammo- 
niacal  salts,  as  the  nitrate  oxalate,  &c.,'^  without 
any  disadvantage  to  the  nutrition  and  to  the  dis- 
appearance of  ammonia  ;  and  thus,  far  from  being  de- 
composed, in  order  to  supply  ammonia  during  its 
development,  and  during  the  fermentation,  as  Dobereiner 
had  asserted,  the  yeast  consumes  the  ammonia  contained 
in  the  liquids  which  are  fermenting. 

Although  a  salt  of  ammonia  may  serve  for  the 
nutrition  and  development  of  ferment,  it  is  proved, 
however,  by  generally  observed  facts,  that  it  is  not  the 
especial  nitrogenous  aliment  for  this  simple  organism. 
If  we  substitute  for  the  ammoniacal  salt  in  Pasteur's 
experiment  natural  juices  (as  that  of  grapes,  or  of 
beetroot,  or  the  water  in  which  yeast  has  been  washed), 
containing  nitrogenous  organic  substances,  the  quantity 
of  yeast  formed  and  decomposed  in  the  same  time 

*  In  all  the  observations  made  by  Pasteur  and  the  other  experimentalists, 
an  increase  or  decrease  of  the  rate  of  development  of  the  cells,  their  multipli- 
cation and  their  nutrition,  was  always  accompanied  by  a  variation  in  some 
direction  in  the  energy  with  which  the  sugar  (one  of  the  essential  nutritive 
elements)  was  resolved  into  alcohol  and  carbon  dioxide.  W^e  will  discuss 
presently  what  explanation  may  be  given  of  the  correlation  between  the  two 
phenomena. 


84 


ON  FERMENTATION. 


is  much  greater,  and  the  decomposition  of  sugar  is 
more  active. 

There  exist,  therefore,  nitrogenous  carburetted  sub- 
stances which  are  better  suited  to  the  nutrifion  of 
yeast  than  ammonia.    What  are  they? 

The  natural  juices  which  we  have  just  mentioned, 
and  more  especially  yeast-water  which  shows  itself  to 
be  particularly  active,  contain  different  kinds  of  nitro- 
genous matter,  and  particularly  albuminoid  substances. 
Direct  experiments  alone  can  determine  whether  we  are 
to  attribute  the  active  part  played  by  these  juices  to 
proteinic  principles,  or  to  more  simple  compounds. 

Pasteur  found  that  the  albumin  of  the  white  of  egg 
was  entirely  unfit  for  the  nourishment  of  the  globules 
of  yeast.  Even  at  an  earlier  period  M.  Thenard  and 
M.  Colin  had  observed  that  albumin  does  not  begin 
to  excite  alcoholic  fermentation  (or,  which  is  the  same 
thing,  to  produce  the  nutrition  and  development  of 
the  yeast  globules)  till  the  end  of  three  weeks  or  a 
month,  when  left  at  the  temperature  of  30°  C.  (86°  Fahr.), 
when  it  undergoes,  under  the  influence  of  infusoria  and 
mucidines,  which  are  developed  in  it,  a  greater  or 
less  degree  of  decomposition.  The  serum  of  blood 
encourages  the  nutrition  of  the  globules,  without  re- 
quiring to  be  itself  previously  decomposed  ;  but  it  is 
not  the  serine  which  is  active  in  this  case ;  for,  if 
we  eleminate  it  by  coagulation,  the  boiled  and  filtered 
liquid,  when  sugar  and  yeast  are  added,  rapidly 
produces  an  energetic  fermentation  (Pasteur). 

M.  Mayer  has  made  many  experiments,  with  the 
view  of  throwing  light  on  the  question  of  the  nutritive 
part  played  by  albuminoid  substances.    He  has  found 


FUNCTIONS  OF  YEAST. 


85 


that  the  inactivity  of  the  greater  part  of  them  (albumin, 
casein,  &c.)  arises  especially  from  their  not  being 
diffusible  through  the  organized  membranes  of  the 
cells.  We  know,  in  fact,  that  most  of  these  bodies 
belong  to  the  class  of  colloid  substances,  not  diffusible 
through  porous  membranes,  and  we  can  easily  under- 
stand that,  not  being  able  to  penetrate  into  the  interior 
of  the  cell,  but  remaining  imprisoned  in  the  surrounding 
liquid,  they  will  not  be  in  a  condition  to  give  any 
powerful  aid  to  the  development,  the  nutrition,  and 
the  multiplication  of  the  globules.  The  diffusible 
products  formed  by  stomachic,  intestinal,  or  artificial 
digestion,  show  themselves  to  be  eminently  suited  to 
nourish  the  cell  of  the  Saccharomyces, 

It  is  the  same  with  regard  to  diastase,  and  the 
different  kinds  of  pepsin  ;  but,  as  the  nutritive  activity 
of  these  ferments  remain  after  they  are  cooked,  it 
must  by  no  means  be  attributed  to  their  specific 
properties  as  soluble  ferments  (for  these  properties 
are  destroyed  by  heat),  but  rather  to  products  analogous 
to  peptone,  which  always  accompany  these  soluble 
ferments,  and  of  which  it  is  very  difficult  to  get  rid 
— syntonin,  and,  in  a  less  degree,  allantoin,  urea 
guanine,  and  uric  acid,  increase  the  fermenting 
power  of  yeast ;  that  is  to  say,  give  nourishment  to 
the  organism. 

Other  nitrogenous  substances,  which  we  ought  also 
to  consider  as  compound  ammonias,  have  shown 
themselves  to  be  but  slightly,  or  not  at  all,  active. 
Such  are  creatin,  creatinine,  caffeine,  asparagin,  leucine, 
hydroxylamine. 

It  is,  then,  very  probable,  according  to  these  results, 


86 


ON  FERMENTATION. 


that  natural  juices,  the  wort  of  beer,  and  water  in 
which  fresh  yeast  has  been  washed,  owe  their  power 
of  nourishing  the  cells  of  yeast,  not  to  albuminoid 
principles,  properly  so  called,  which  are  indififusible, 
but  to  allied  nitrogenous  compounds  analogous  to  pep- 
tones, which  have  the  property  of  passing  by  osmose 
through  membranes. 

Yeast  thus  affords  us  an  evident  and  striking  example 
of  vegetable  cells,  which  assimilate  their  nitrogen  under 
the  form  of  complex  combinations,  allied  by  their  con- 
stitution to  the  higher  forms  of  albuminoid  substances. 
There  is  no  proof  that  similar  phenomena  are  not  pro- 
duced in  plants  of  higher  organization. 

Agricultural  and  physiological  experiments  do  not 
establish  so  clearly  the  assimilation  of  nitrogenous 
organic  combinations  as  that  of  nitrates  and  ammo- 
niacal  salts ;  although  the  few  observations  made  on 
this  subject  are  rather  favourable  than  otherwise  to 
a  positive  solution  of  the  question.  But,  if  it  were 
otherwise,  it  would  not  be  logical  and  prudent  to  seek 
to  draw  a  sharp  distinction  between  the  phenomena 
of  nutrition  of  the  Sacckaromyces,  and  those  of  larger 
plants,  and  to  say  that  the  former  may  find  nitrogenous 
nourishment  in  the  organic  nitrogenous  combinations 
allied  to  albuminous  substances,  and  that  the  latter  do 
not. 

Plants  of  complex  organization  are,  in  fact,  formed 
by  the  union  of  cellular  elements  of  various  kinds, 
fulfilling  different  functions,  and  whose  conditions  of 
nutrition  and  development  are  not  identical ;  among 
which  it  is  probable  that  some  are  to  be  found 
susceptible   of  assimilating  complex   nitrogenous  or- 


FUNCTIONS  OF  YEAST. 


87 


ganic  materials,  that  have  been  elaborated  elsewhere 
at  the  expense  of  ammoniacal  salts,  or  of  nitrates. 

Assimilation  of  Mineral  Principles. — Vegetables,  in 
general,  always  leave,  after  combustion  in  air  or  in 
oxygen,  a  fixed  mineral  residuum,  whose  weight  varies, 
within  certain  limits,  from  one  vegetable  species  to 
another,  and  also,  especially,  from  one  organ  to  another, 
as  well  as  for  the  same  organ,  according  to  its  age. 
Thus,  the  leaves  of  the  pear-tree  yielded  to  M.  Violette, 
7*118  grammes  of  ash  per  cent,  of  dried  matter  : — 


The  extremity  of  the  twigs 
The  middle  part . 
The  lower  part  •  • 
The  trunk        •  • 
The  roots         •  • 


jbark  . 

I  wood  . 

( bark  • 

(wood  . 

jbark  . 

I  wood  . 

jbark  . 

(wood  , 

{bark  • 

wood  • 


3*454 
0-304 

3*682 
0-134 
2-903 
o*354 
2-657 
0*296 

1*127 
0-234 


In  proportion  as  the  plant  grows  older,  the  weight  of 
the  ash  increases. 

Do,  then,  mineral  principles,  which  are  found  in  all 
vegetables,  from  the  highest  point  of  the  scale  to  the 
lowest — for  we  have  already  seen,  by  analysis,  that  yeast 
forms  no  exception — do,  then,  mineral  principles  play 
an  important  part  in  the  biological  phenomena  of  the 
nutrition  and  development  of  the  plant,  or  do  they 
only  make  their  appearances  as  useless,  but  not 
injurious  elements,  inevitably  introduced  by  the  fluids 
from  which  the  plant  derives  its  nutritive  principles  ? 


88 


ON  FERMENTATION. 


The  remarkable  constancy  of  the  chemical  composi- 
tion of  various  kinds  of  ash,  especially  with  respect  to 
their  constituent  elements,  as  well  as  the  most  complete 
agricultural  experiments,  have  proved  in  the  most  posi- 
tive manner  that  the  greater  part  of  saline  compounds, 
found  by  analysis,  are  necessary  to  vegetation.  We 
have  also  learned  to  classify  them  in  the  order  of  their 
nutritive  importance,  with  reference  to  the  entire  plant 
and  its  different  constituent  parts  (leaves,  stalks,  seeds, - 
grain,  &c.). 

Thus  the  phosphates  preponderate  remarkably  in 
grain,  and  form  by  themselves  the  whole  of  the  mineral 
mass  found  after  incineration,  as  shown  in  the  following 
results,  published  by  Berthier  :— 


Phosphates  in  too  Parts  of  Ash. 


Nature  of  Phosphate. 

White  Wheat— 
Chartrain. 

Rye. 

Barley. 

0 

1        Rice  from 
[  Carmargue. 

Maize. 

French  Beans— 
Soissons. 

Green  Peas. 

Lentils. 

Potassium  phosphate 

50 'CO 

48-50 

52*50 

7 '50 

24*10 

41*50 

42*70 

66*70 

61*70 

Calcium 

22 '00 

29*20 

15-00 

16-50 

24*10 

18-50 

8*40 

22 '20 

6*50 

Magnesium 

28  "oo 

25-00 

20-00 

24-10 

38-00 

14*30 

6 -60 

19*60 

Manganese  „ 

18-30 

Total   

1 00  "go 

96*00 

92-50 

44*00 

72-30 

65*50 

98*09 

95*50 

87*80 

Wc  also  give,  for  reference,  from  the  same  author, 
the  analyses  of  the  ashes  of  various  parts  of  plants. 


FUNCTIONS  OF  YEAST.  89 


Stalks,  icq  Parts  of  Ash  contain— 


0  3 

c 

Com  osition  of  the  Ash-s 
omposi  ion  0      e    s  .v,s. 

Vine 
Nemo 

Whe 
Stra 

Luce 

r^otasn  ,,,       ,,.  ... 

3*40 

_ 

Potassium  and  sodium  car-^ 
bonates    j 

16*40 

14-44 

12 '20 

Potassium  chloride 

2 '20 

0 

78 

2*50 

1*90 

3  04 

sulphate  ... 

4-40 

3 

•40 

030 

2-66 

1-30 

phosphate 

silicate  .,, 

Lime  ... 

4 

00 

1570 

Calcium  carbonate   

49-82 

6 

CO 

64 '26 

22*62 

Magnesium    , ,   

3-85 

6*07 

6*39 

Carbon  dioxide   

I 

00 

Iron  oxide   

2 '60 

Calcium  phosphate   

1570 

6 

60 

9'oo 

8-43 

11*31 

Magnesium   

Iron  ,,   

i"83 

Manganese  ,,   

Phosphoric  acid 

I '20 

Silica  

5-80 

78 

•22 

73 '90 

2 '24 

39-80 

Bulbs  and  Roots  gave,  for  ioo  Parts  Ash — 


Madder 
Root. 

Jerusalem 
Artichoke. 

Potatoes. 

Onions. 

Potassium  and  sodium  car-) 

31  "50 

42-43 

21*60 

bonates         ...       ...  j 

31*11 

Potassium  chloride   

3*14 

7 '50 

4*oo 

2*20 

Sodium          , ,   

Potassium  sulphate   

3  93 

6-00 

2*8o 

4*00 

phosphate  

Calcium  carbonate   

30  00 

3470 

35 'oi 

2-80 

12*00 

Magnesium    , ,   

4-13 

10*00 

Calcium  phosphate   

9-71 

16*^50 

6*87 

38*00 

Magnesium  ,,   

8-50 

2*50 

Tron   

5 '09 

1*70 

Silica   

7*88 

2*50 

5 


90 


ON  FERMENTATION. 


In  the  leaves,  the  calcium  carbonate  and  the  silica 
preponderate,  and  form  in  themselves  from  60  to  90  per 
cent,  of  the  total  weic^ht  of  ash. 


Pine  Leaves. 

Vine  Leaves. 

Mulberry- 
Leaves. 

Calcium  carbonate  

6874 

51-00 

53 'OO 

Silica  ..r   

6*43 

10-20 

27*70 

Total   

75'i7 

61-20 

80-70 

Let  us  compare  with  this  the  composition  of  the  ash 
of  beer-yeast  (Mitscherlich)  : — 


Phosphoric  acid 
Potassa  . 
Soda 

Magnesium  phosphate 
Calcium  „ 


Surface  Yeast. 
.  41-8  . 
.    39-8  . 

.  1 6-8 
.  2*3 


Sedimentary  Yeast. 

39*5 
28-5 

.  22*6 

97 


If  mineral  salts  really  play  an  active  part  in  vegeta- 
tion, we  may  foresee,  from  these  analyses, — 

First.  That  their  relative  importance  will  vary  with 
the  respective  weights  of  these  different  bodies,  found 
in  organized  tissues  ;  that,  in  consequence,  the  phos- 
phates, potash,  soda,  magnesia,  and  the  sulphates  will 
occupy  the  first  place. 

Secondly.  That  according  as  the  soil  or  the  medium 
in  which  the  plants  grow  is  more  especially  manured  by 


FUNCTIONS  OF  YEAST. 


91 


any  one  of  these  salts  in  particular,  the  development 
either  of  leaves,  stalks,  or  seeds  will  be  favoured.  These 
results  have  all  been  verified  by  direct  experiment,  but 
as  it  does  not  enter  into  our  plan  to  study  agricultural 
chemistry  with  reference  to  mineral  manures,  and  as 
our  intention  is  merely  to  compare  the  nutrition  of 
yeast,  and  of  analogous  organisms,  with  that  of  other 
plants  of  a  higher  order,  we  will  return  to  the  history  of 
the  Saccharomyces.  We  would  remark  that  the  compo- 
sition of  its  ash,  entirely  composed  of  phosphates, 
approaches  more  nearly  to  that  of  seeds,  to  which  it 
is  also  allied  by  the  analogy  of  function  and  general 
chemical  composition. 

We  owe  to  M.  Pasteur  the  proof  of  the  absolute 
necessity  of  mineral  salts  (phosphates  of  the  alkalis, 
and  the  alkaline  earths)  for  the  development  and 
nutrition  of  the  yeast-cell.  If,  in  his  experiment,  in 
which  the  ferment  is  sown,  in  an  imponderable  quantity, 
in  a  medium  entirely  composed  of  pure  sugar-candy, 
ammonium  tartrate,  and  ash  of  yeast,  we  omit  the 
latter  element,  the  fermentation  and  development  of 
cells  which  ought  to  precede  it  {i.e.^  the  fermentation), 
do  not  take  place.  M.  Pasteur  went  no  farther  in  these 
researches  ;  absorbed  by  the  pursuit  of  a  different  aim, 
he  did  not  endeavour  to  ascertain  what  were  the  most 
favourable  mineral  substances ;  he  only  used  in  his  re- 
searches ash  of  fresh  yeast  as  an  inorganic  element, 
rightly  thinking  that,  at  all  events,  he  should  find  in  it 
that  which  agrees  best  with  the  mineral  nutrition  of  the 
fungus. 

M.  Mayer,  following  up  this  work,  and  seeking  to 
ascertain  by  direct  experiment  v/hich  among  the  salts 


92 


ON  FERMENTATION. 


generally  contained  in  varying  proportions  in  vegetable 
ashes  clearly  favour  the  development  of  yeast,  arrived 
(loc,  cit)  at  the  following  conclusions  : — 

1.  Preparations  of  iron,  employed  in  very  small  quan- 
tities, seem  to  have  no  influence ;  in  larger  proportions 
they  are  injurious. 

2.  Potassium  phosphate  shows  a  preponderating 
favourable  influence.  It  may  be  employed  in  a  liquid 
medium,  in  a  high  percentage,  without  its  fertilizing 
influence  being  destroyed  ;  while,  for  plants  of  a  higher 
order,  so  great  a  concentration  would  become  a  serious 
cause  of  pathological  disturbance.  Potassium  phos- 
phate is  not  only  favourable,  but  indispensable. 

In  fact,  if  from  a  medium  formed  of  sugar-candy, 
ammonium  nitrate,  traces  of  yeast,  and  a  mixture  of 
acid  potassium  phosphate,  magnesium  sulphate,  and 
tricalcic  phosphate,  a  medium  which  ferments  with 
considerable  activity,  we  omit  the  acid  potassium 
phosphate,  fermentation  and  the  development  of  yeast 
are  not  produced. 

Potassium  phosphate  can  by  no  means  be  replaced  by 
sodium  phosphate,  which  is  inactive. 

The  absence  of  calcium  phosphate  from  the  medium 
causes  much  less  injurious  consequences  than  the 
omission  of  the  before-mentioned  salt. 

The  result  is,  that  potassium  and  phosphoric  acid  are 
indispensable  elements,  whilst  lime  may  be  omitted 
without  any  great  inconvenience,  as  we  might  have  fore- 
seen from  the  results  of  the  analysis. 

Magnesium,  on  the  contrary,  appeared  in  Mayer's 
experiments  to  be  a  very  useful,  if  not  an  indispensable, 
clement.  It  is  immaterial  whether  this  metal  is  supplied 


FUNCTIONS  OF  YEAST. 


93 


under  the  form  of  sulphate,  or  of  ammoniaco-magnesian 
phosphate. 

The  combinations  of  sodium  present  no  material 
effects,  conformably  with  what  has  been  already  ob- 
served in  plants  of  a  higher  order. 

The  sulphur,  administered  to  yeast  under  the  form  of 
sulphates,  or  soluble  sulphites,  appears  not  to  be  assimi- 
lated. At  least,  the  presence  or  absence  of  these  two 
classes  of  salts  seems  to  have  no  influence.  Yet  yeast 
contains  sulphur  in  appreciable  proportions,  which  w^e 
even  find  combined  intimately  in  the  products  of  its 
dis-assimilation  (sulphuretted  pseudo-leucine  of  Heintz). 
We  are  unable  to  say  what  the  origin  of  this  normal 
sulphur  may  be. 

M.  Raulin,  in  a  remarkable  investigation,  has  studied 
with  particular  care,  and  by  an  excellent  method,  the 
influence  of  the  mineral  components  of  the  medium  on 
the  development  of  a  cellular  plant,  the  Aspergillus  niger. 
As  the  results  obtained  may  be  interesting  with  respect 
to  the  question,  rather  as  a  general  one  than  specially 
relating  to  yeast,  which  we  are  now  considering,  we  will 
enter  into  some  details  on  this  point,  more  especially 
since  the  experimental  method  employed  by  M.  Raulin 
may  perhaps  serve  as  a  model  for  other  researches  of 
this  kind. 

First  of  all,  an  artificial  medium  is  prepared,  ex- 
clusively formed  of  definite  chemical  compounds  suitable 
for  the  vegetation  of  a  particular  plant.  In  order  to  study 
the  influence  of  various  physical  or  chemical  circum- 
stances on  the  development  of  this  plant,  a  vessel  is 
filled  with  the  artificial  mixture  and  placed  under  the 
most  favourable  conditions  for  its  vegetation.   The  seeds 


94 


ON  FERMENTATION. 


of  the  plant  are  sown  in  it,  and  they  are  allowed  to  grow 
during  the  necessary  time ;  this  trial,  which  is  repro- 
duced exactly  in  the  same  manner  in  each  series  of 
experiments,  is  the  typical  trial,  with  which  all  others 
are  compared. 

Another  experiment  is  arranged  in  every  respect  like 
the  first,  with  the  exception  of  the  single  circumstance 
which  it  is  proposed  to  study.  The  two  crops,  obtained 
at  the  same  time,  are  dried  and  weighed  sepai:ately,  and 
the  numerical  ratio  of  the  weight  of  these  two  results 
will  be  the  measure  of  the  influence  of  the  condition  to 
be  examined. 

The  degree  of  perfection  of  the  method  depends  upon 
three  general  conditions  : 

1.  It  is  absolutely  necessary  to  find,  in  the  first  place, 
an  artificial  medium  suited  to  the  development  of  the 
plant  to  be  studied.  M.  Raulin  found  the  ground  quite 
prepared  in  this  respect,  thanks  to  the  labours  of 
M.  Pasteur :  the  latter  had  observed  that  the  mucidines 
{Penicillitirn)  can  be  developed  in  a  medium  exclusively 
formed  of  definite  artificial  substances. 

Water,  sugar,  ammoniacal  salt  (bitartrate),  and  ash 
of  yeast.  No  portion  whatever  of  the  constituent  parts 
of  this  medium  can  be  omitted,  without  giving  a  com- 
plete check  to  the  development. 

2.  The  weight  of  the  crop  which  the  medium  intended 
for  the  typical  experiments  can  yield  in  a  given  time, 
with  a  constant  weight  of  nutritive  substances,  ought,  all 
other  things  being  equal,  to  be  as  great  as  possible. 

3.  The  typical  experiments  placed  under  the  same 
conditions  ought  to  yield  crops  whose  numerical  ratios 
differ  but  little  from  that  assumed  as  unity :  the  ratio 


FUNCTIONS  OF  YEAST. 


95 


which  differs  the  most  fixes  the  relative  maximum  error 
of  the  process. 

At  the  beginning  of  his  researches,  M.  Rauhn,  rely- 
ing on  the  data  of  M.  Pasteur,  made  use  of  a  typical 
medium  composed  of — 


Water 

Sugar       .      .  • 
Ammonium  nitrate  • 
Tartaric  acid 
Ammonium  phosphate 
Potassium  carbonate 
Calcium  „ 
Magnesium  „ 
Aspergillus  sowed  at  20°  C. 


2,000 
70 

3 

2 


small 
C  quantities. 

Fahr.). 


With  such  a  medium,  the  variation  in  weight  of  the 
crop,  between  one  typical  experiment  and  another,  was 
so  considerable,  that  it  was  not  possible  to  ascertain  the 
influence  exercised  by  the  omission  of  certain  elements, 
which  entered  into  the  mixture  only  in  small  propor- 
tions. Thus,  after  forty-eight  hours*  vegetation,  the 
weight  of  two  typical  crops  were  found  equal  to — 


No.  I 


Grammes. 

.  3*i9- 


No.  2  , 


Grammes. 
,  177. 


By  the  omission  of  all  the  mineral  elements  he  arrived 
at  the  result — 

Grammes.  Grammes. 
No.  I  .    .    .    o'lo.  No.  2  .    .    .  0*87. 


The  omission  of  potassium  carbonate' alone  gave — 


Grammes. 

No.  I  .   .   .  2*29. 


Grammes. 
No.  2  .   •   .  I'll. 


96 


ON  FERMENTATION. 


The  action  of  the  whole  of  the  mineral  salts  comes 
out  strongly ;  that  of  the  potassium  carbonate  is  not 
perceptible,  for  the  number  2*29  lies  between  3*19  and 
1*177,  the  numbers  found  for  the  typical  experiments. 

Besides,  in  these  first  experiments,  M.  Raulin  ascer- 
tained that  the  development  of  the  mucidines  was 
fairly  rapid  in  the  first  few  days,  and  then  grew  indefi- 
nitely slower.  While  seeking  to  find  the  causes  of  this 
disturbance  by  tentatively  modifying  the  conditions  of 
the  medium,  especially  by  adding  to  it  sulphur,  zinc, 
iron,  and  silicon,  in  the  form  of  salts  ;  by  modifying  the 
proportions  of  the  essential  elements,  raising  the  tem- 
perature to  35°  C.  (95°  F.)  ;  and,  finally,  by  employing 
vessels  of  considerable  area  and  small  depth,  he  suc- 
ceeded in  finding  a  typical  medium,  giving  for  the  same 
length  of  time  a  result  fifty  times  greater  than  that  of 
the  first  experiments.  Under  these  conditions,  the  ratio 
of  the  typical  experiments,  instead  of  varying  from  I'D 
to  1*8,  acquired  a  remarkable  constancy,  and  did  not 
vary  now  more  than  of  its  value.  It  is  evident  that 
the  favourable  influence  of  any  particular  substance  will 
then  show  itself  in  a  much  more  defined  manner. 

The  experiments,  as  far  as  Aspergillus  niger  is  con- 
cerned, must  now  be  conducted  in  the  following 
manner : — 

In  the  vessel  intended  for  the  typical  experiment,  the 
following  chemical  substances  are  brought  together : — 


Water    ......  1,500 

Sugar-candy  70 

Tartaric  acid  .       .       .       •       •  4 
Ammonium  nitrate        ...  4 
„        phosphate  .      •      .  o'6o 


FUNCTIONS  OF  YEAST. 


97 


Potassium  carbonate  •      .       •  o"6o 

Magnesium       „  ...  0*40 

Ammonium  sulphate  .       .       .  0*25 

Zinc               „  ...  0-07 

Iron               „  ...  0*07 

Potassium  silicate  .       .       .  0*07 


This  mixture  is  left  to  itself  for  several  hours,  and 
then  stirred  with  a  porcelain  spatula. 

In  order  to  sow  the  fungus,  it  is  sufficient  to  pass  over 
the  whole  surface  the  end  of  a  camels'-hair  brush  with 
which  spores  have  been  collected  from  a  very  pure,  and 
not  too  dry,  vegetation  of  Aspergilhis, 

When  we  are  not  yet  in  possession  of  any  Aspergillus, 
it  is  sufficient,  in  order  to  procure  this  plant  in  a  pure 
state,  to  leave  exposed  to  the  air  certain  natural  sub- 
stances, such  as  the  water  of  acidulated  yeast,  damp 
bread,  or  slices  of  lemon.  The  spores  of  Aspergillus 
which  exist  among  the  germs  in  the  atmosphere  may 
fall  on  these  matters,  and  develop  themselves  there, 
mixed  with  other  organisms. 

When  we  see  Aspergillus  make  its  appearance,  which 
is  immediately  distinguishable  by  its  black  fructification, 
it  is  again  sown  on  an  artificial  liquid,  and  we  at  last 
obtain  it  free  from  mixture.  The  typical  experiment 
being  thus  prepared,  it  is  placed  in  a  stove  at  35°  C. 
(95°  F.),  and  constantly  supplied  with  damp  air.  The 
spores  develop,  and  at  the  end  of  twenty-four  hours  the 
filaments  of  the  mycelium  form  a  continuous  whitish 
membrane  on  the  surface  of  the  liquid  At  the  end  of 
forty-eight  hours  this  membxane  has  become  very  thick, 
and  turns  to  a  deep  brown  colour;  after  three  days  it  has 
become  quite  black  on  the  upper  surface,  which  colour  is 


98 


ON  FERMENTATION. 


due  to  the  appearance  of  spores.  The  thick  membrane 
is  then  removed  by  the  fingers,  squeezed,  and  then  spread 
upon  a  plate  to  dry.  New  spores  are  sown  upon  the 
Hquid,  and  after  three  days  we  obtain  a  second  crop, 
weaker  than  the  first. 

The  typical  mixture,  and  that  which  is  to  serve  for 
the  experiment,  and  which  differ  from  each  other  only 
in  the  single  element,  the  influence  of  which  we  wish  to 
ascertain,  are  placed  together  in  the  stove.  This  in- 
fluence is  measured  by  the  ratio  of  the  weights  of  the 
two  first  crops,  or,  still  better,  by  the  amount  of  the 
first  and  second  crops  obtained  in  six  days. 

Example. — The  following  trial  mixtures  were  placed 
in  the  stove : —  . 

No.  I.  Typical  medium. 

No.  2.        „  less  the  potass. 


Ratio  of  the  two  first  crops  =  j -g  ;  that  of  the 
weights  of  the  whole  crops  numbers  which  prove 

plainly,  in  the  most  complete  manner,  the  utility  of  the 
potass. 

The  results  obtained  by  this  remarkable  method  are 
the  following: — 

I.  All  the  elements  of  the  typical  artificial  medium 
concur  simultaneously  in  the  development  of  the  plant, 
for  if  we  omit  each  of  them  in  turn,  the  weight  of  the 
crop  undergoes  a  diminution,  which  is  usually  somewhat 


First  crop  (after  3  days)  . 
Second  crop  (after  3  other  days) 


No.  I.  No.  2. 

Grammes.  Grammes. 

,    14*4  0*8o 

,     lO'O  0*I2 


Total  24-4 


0*92 


FUNCTIONS  OF  YEAST. 


99 


considerable,  and  which  cannot  be  attributed  to  experi- 
mental errors. 

2.  The  mineral  oxides  of  the  artificial  medium  cannot 
be  substituted  for  each  other. 

3.  Nitric  acid  may  be  used  instead  of  the  ammoniacal 
salts  as  the  nitrogenous  aliment. 

Finally,  the  following  are  the  ratios  found  between 
the  typical  and  the  experimental  trials  : — 


Omission  of  the  oxygen    .      *       •      .  very  great 

„  „  „  water      .       .       *       .  infinite 

„  „  „  sugar      .       .       .       .       .  65 

„  „  tartaric  acid  ....  infinite 

„  „  ammoniacal  salt,  or  nitrate  .       •    1 53 

„  „  phosphoric  acid    .       .       .  .182 

„  „  magnesia  91 

5,  „  potash  25 

„  „  sulphuric  acid       ,       .       .  .24 

„  „  zinc  oxide  10 

„  „  iron  oxide     .       .      .      .      .  27 

„  „  siHca  1*4 


The  nutritive  elements  of  the  artificial  medium  are 
some  indispensable,  which  are  those  found  in  large  pro- 
portions in  it,  and  others  are  useful,  but  apparently  not 
indispensable  :  these  only  enter  into  the  composition  of 
the  typical  medium  in  very  small  proportions. 

It  is  probable  that  some  of  these,  such  as  sulphur, 
exist  accidentally  in  very  small  quantities  in  the  artificial 
media,  to  which  they  have  not  been  added,  and  may 
thus  set  up  a  sluggish  development.  Thus,  Mayer 
asserts  that  he  has  been  unable  by  repeated  crystal- 
lizations, and  even  by  precipitating  the  liquor  by  barium 
chloride,  to  obtain  sugar  free  from  sulphur ;  it  is  to  the 


lOO 


ON  FERMENTATION. 


presence  of  this  sulphur  that  he  attributes  the  introduc- 
tion of  this  element  into  the  newly  formed  yeast. 

It  is  evident  that  the  results  obtained  by  M.  Raulin, 
and  especially  his  method,  may  be  applied  to  the  search 
after  better  conditions  for  the  maximum  development 
of  other  kinds  of  vegetation,  or  simple  organisms.  M. 
Pasteur,  who  in  his  laboratory  at  the  "  Ecole  normal " 
discovered  new  methods  for  the  production  of  pure  beer- 
yeast  on  a  large  scale,  had  to  make  preparatory  trials 
analogous  to  those  of  M.  Raulin.  His  researches  also 
establish  an  important  fact,  that  an  artificial  medium, 
suitably  prepared,  may  be  as  favourable  to  the  develop- 
ment of  vegetation,  and  even  more  favourable,  than 
the  most  fertile  natural  media.  We  may  thence  draw 
conclusions  of  great  importance  as  to  the  cultivation  of 
larger  plants,  and  may  suppose  that  chemical  manures, 
suitably  chosen,  may  be  substituted  for  natural  ones  in 
agriculture,  with  great  advantage. 

This  is  what  several  men  of  science,  who  make  a 
study  of  agriculture,  have  already  attempted  ;  the  great 
point  is  to  determine  carefully  the  useful  composition  of 
these  manures  ;  unfortunately,  we  must  admit,  experi- 
ments on  larger  plants  are  not  so  simple  and  so  easily 
managed  as  those  on  mucidines. 

Sugar. — Pasteur  and  Raulin  have  demonstrated  the 
preponderating  influence  of,  and  the  necessary  part 
played  by,  sugar  or  analogous  bodies  in  the  vegetation 
of  Aspergillus  and  of  mucidines.  This  influence  is  as 
powerful  in  the  development  of  the  yeast  of  beer. 
Without  sugar,  without  hydrocarbonate  substances, 
yeast  can  neither  reproduce  nor  be  nourished.  An 
important  difference  is  thus,  at  first  sight,  established 


FUNCTIONS  OF  YEAST. 


lOI 


between  simple  organisms,  such  as  ferments,  mildews, 
&c.,  and  the  larger  plants,  which  derive  the  organic 
elements  of  their  constitution  from  the  simplest  com- 
pounds of  carbon,  such  as  carbon  dioxide. 

This  distinction,  however,  loses  its  force  after  a  more 
complete  examination. 

If  the  larger  plants  derive  nourishment  at  the  expense 
of  carbon  dioxide,  it  is  because,  in  their  leaves  and  other 
green  parts,  there  are  found  organs  suited  to  the  utiliza- 
tion of  the  active  force  of  the  luminous  rays  sent  by 
the  sun  or  other  source  of  light.  The  carbon  is  set  free 
directly,  and  the  oxygen  is  disengaged.  It  appears  very 
probable,  that  at  the  moment  when  the  carbon  is  sepa- 
rated from  the  oxygen,  under  a  special  condition  as 
yet  unknown,  and  very  different  from  that  of  black 
amorphous  carbon,  or  of  the  diamond  or  graphite  (forms 
under  which  we  know  this  element),  that  it  unites  with 
the  elements  of  water  to  form  a  hydrate  of  carbon 
(starch,  sugar  ?),  or  at  least  a  body  which  can  be  converted 
into  these  principles  by  ulterior  transformations. 

If  we  were  able  to  effect  the  decomposition  of  carbon 
dioxide  under  the  influence  of  light  outside  the  animal 
economy,  I  have  no  doubt  but  that  (if  the  experiment 
were  made  in  the  presence  of  water)  there  would  be 
found  a  hydrocarbon  compound.  I  have  even  been 
able  to  give  a  slight  experimental  confirmation  of  this 
theoretical  opinion. 

If  we  treat,  in  the  cold,  coarsely  powdered  white  cast 
iron  (which  is  known  to  contain  an  iron  carburet),  with 
a  solution  of  cupric  sulphate,  the  iron  of  the  white  cast 
iron  is  entirely  dissolved,  without  disengagement  of  any 
carbon  or  other  gas  ;  after  having  washed  it,  we  may 


102 


ON  FERMENTATION. 


eliminate  the  deposited  copper  by  placing  it  in  contact 
with  a  solution  of  iron  perchloride.  •  The  copper  is 
rapidly  dissolved ;  there  remains  a  pulverulent  black 
mass,  which,  after  dessication  at  80°  C.  (176°  Fahr.),  in  a 
vacuum,  resembles  carbon.  But  this  carbon  contains 
w^ater  in  combination,  which  is  suddenly  disengaged  when 
it  is  heated  to  about  250°  C.  (480°  Fahr.)  ;  it  is  easily  dis- 
solved in  nitric  acid,  becoming  oxidated,  yielding  yellow 
or  orange-yellow  substances  containing  nitrogen.  This 
residuum,  when  analyzed,  gives  a  quantity  of  water, 
which  is  in  a  tolerably  constant  proportion  to  that  of 
carbon. 

It  therefore  represents  a  true  and  definite  carbon 
hydrate.  It  is  evident  that  the  condition  of  the  carbon 
in  the  cast  iron  must  be  very  different  from  that  of  the 
carbon  of  carbon  dioxide,  and  that  the  hydrates  which 
arise  from  the  separation  of  these  forms  of  carbon  may 
differ  greatly.  Nevertheless,  the  experiment  which  I 
have  just  described  gives  material  support  to  the  idea 
which  physiologists  entertain  of  the  successive  chemical 
metamorphoses  of  the  carbon  compounds  in  plants. 

When  the  carbon  hydrate  is  once  formed  in  the  leaf, 
it  is  carried  into  the  other  parts  of  the  plant,  to  serve 
there  as  nutrition,  for  the  development  of  cells  con- 
taining no  chlorophyll,  and  whose  biological  functions 
closely  resemble  those  of  cellular  organisms.  That 
which  takes  place  during  the  germination  of  seeds,  up 
to  the  moment  when  the  new  plant  becomes  provided 
with  aerial  leaves  which  have  become  green  under  the 
influence  of  light  and  air,  and  begins  to  utilize  carbon 
dioxide,  leaves  no  doubt  as  to  the  scientific  value  of  this 
interpretation. 


FUNCTIONS  OF  YEAST. 


103 


We  see  here  the  newly  formed  cells  successively 
developed,  and  superposed  so  as  to  form  radicles,  stalk, 
cotyledons,  and  leaves,  and  the  germ  procures  the  neces- 
sary materials  for  its  development  from  the  organic 
principles  which  the  seed  has  accumulated,  and  among 
which  hydrocarbons  are  always  predominant. 

We  must,  therefore,  admit  that  the  phenomena  of 
nutrition  of  the  larger  plants  do  not  seem  to  differ 
much,  when  examined  in  detail,  from  those  of  the  more 
simple  ones. 

The  former  are  provided  with  special  organs  which 
enable  them  to  elaborate  for  themselves  the  hydro- 
carbon substances  which  they  require  for  the  develop- 
ment of  the  rest  of  their  organism.  The  inferior 
cellular  plants,  and  in  fact  generally  all  those  which  are 
unprovided  with  cells  containing  chlorophyll,  are  neces- 
sarily parasites,  which  must  borrow  their  hydrocar- 
bonate  nourishment,  directly  or  indirectly,  from  plants 
furnished  with  these  cells. 

Besides  these  general  considerations,  founded  on  the 
phenomena  of  nutrition  observed  in  plants,  the  experi- 
ments of  M.  Pasteur  establish  with  certainty,  that  in  all 
alcoholic  fermentation  a  part  of  the  sugar  is  fixed  in 
the  yeast,  in  the  state  of  cellulose  or  some  analogous 
body.  In  fact,  since  infinitely  small  quantities  of  yeast, 
sown  in  a  medium  entirely  formed  of  pure  sugar-candy, 
of  ammonium  tartrate  or  nitrate  (Mayer),  and  ash  of 
yeast,  develop  and  give  rise  to  very  ponderable  pro- 
portions of  yeast,  considerably  greater  than  the  original 
quantities,  it  cannot  be  doubted  that  the  hydrocarbon 
principles  of  this  new  vegetation  (cellulose,  &c.),  arc 
furnished  by  the  elements  of  the  sugar. 


104 


ON  FERMENTATION. 


The  following  experiments  lead  to  the  same  result*: — - 

M.  Pasteur  submitted  to  fermentation  lOO  grammes 
of  sugar,  about  750  cubic  centimetres  of  water,  2*626  of 
yeast  (weight  of  the  dried  matter). 

After  the  fermentation,  which  lasted  twenty  days,  he 
collected  2*965  grammes  of  yeast  (dried  matter). 

He  likewise  boiled  for  six  or  eight  hours  a  determined 
weight  of  fermented  yeast,  and  also  of  the  same  yeast 
before  fermentation,  with  sulphuric  acid,  diluted  with 
twenty  times  its  weight  of  water  (fermented  yeast  1707 
grammes,  and  unfermented  yeast  1*73  grammes,  dried  at 
a  temperature  of  100°  C.  (212°  F.). 

The  insoluble  residues  were  weighed  on  filters, 
the  weight  of  which  had  been  estimated  ;  they  were 
then  washed,  dried  at  100°  C.  (212°  F.),  and  weighed. 
The  filtered  liquids  were  neutralized  with  barium 
carbonate  ;  the  quantity  of  sugar  formed  by  the  action 
of  the  sulphuric  acid  on  the  cellulose  was  ascertained, 
either  by  means  of  Fehling  s  liquid,  or  by  fermentation. 

He  thus  found,  by  calculating  the  results  obtained  for 
the  two  weights,  2*626  grammes,  and  2*965  grammes  of 
yeast  employed,  and  yeast  obtained — 

1.  That  the  2*626  grammes  of  crude  yeast  employed 
gave  an  insoluble  nitrogenous  residuum  equal  to  0*391 
(14*8  per  cent),  and  0*532  of  fermentable  sugar. 

2.  That  the  2*965  grammes  of  yeast  found  after  fer- 
mentation leave  a  nitrogenizcd  residuum  of  0*634 
grammes  (about  21*4  per  cent),  and  0*918  grammes  of 
fermentable  sugar. 

There  was,  therefore,  fixed  in  the  fermentation  of 
100  grammes  of  sugar  with  2  626  grammes  of  yeast, 
o*4  grammes  of  hydrocarbon  matter,  transformable,  by 


FUNCTIONS  OF  YEAST. 


105 


dilute  sulphuric  acid,  into  fermentable  sugar ;  there 
was  also  a  sensible  augmentation  of  nitrogenous  matter, 
insoluble  in  dilute  sulphuric  acid. 

On  the  other  hand,  in  order  to  verify,  by  a  second 
experiment,  the  value  of  these  conclusions,  M.  Pasteur 
made  use  of  the  process  of  separating  cellulose  from  the 
albuminoid  substances  indicated  by  Payen  and  Schloss- 
berger.  This  process  consists,  as  is  well  known,  in 
treating  yeast  with  dilute  solutions  of  potass. 

In  three  careful  experiments,  M.  Pasteur  found  a 
residuum,  insoluble  in  potass,  formed  of  cellulose,  trans- 
formable into  sugar  by  being  boiled  with  dilute  sul- 
phuric acid  of  1777,  19*29,  and  19*21  per  cent,  of  the 
dry  yeast  experimented  on. 

But  the  0*532  grammes  of  sugar,  produced  without 
the  intervention  of  potass,  by  2*626  grammes  of  the 
same  yeast,  correspond  to  20  per  cent,  of  yeast.  It  is, 
therefore,  proved  that  boiling  in  sulphuric  acid  had 
removed  all  the  cellulose. 

Let  us  also  notice  that  the  2*965  grammes  of  yeast 
found  after  fermentation,  giving  0*918  grammes  of 
sugar,  ought  to  contain  31*9  per  cent,  of  cellulose,  a 
quantity  ii  per  cent,  greater  than  there  was  before 
fermentation.  This  considerable  augmentation  of  the 
weight  of  the  cellulose  in  the  yeast,  while  it  exercised 
its  action  on  the  sugar,  is  a  point  worthy  of  remark, 
since  it  proves  that,  in  accomplishing  one  of  its  principal 
functions,  yeast  undergoes  very  marked  evolutions  in  its 
composition. 

The  following  experiment  of  M.  Pasteur's  proves, 
besides,  that  during  fermentation  the  yeast  itself  forms 
its  fatty  matter  by  the  help  of  the  elements  of  sugar. 


I06  ON  FERMENTATION. 

Let  us  first  call  to  mind  that  Payen's  analyses  shpw 
2  per  cent  of  fatty  matter  in  the  yeast,  and  that  the  lees 
of  wine  also  contain  fatty  matter.  It  had  been  thought 
that  this  fatty  matter  was  furnished  by  the  fermentable 
medium.  Pasteur  mixed  sweetened  water  (prepared 
with  pure  sugar-candy)  with  the  watery  extract  of 
limpid  yeast,  heated  several  times  with  alcohol  and 
ether.  He  sowed  in  it  an  imponderable  quantity  of 
fresh  globules.  These  multiplied,  and  caused  the  sugar 
to  ferment.  He  succeeded  thus  in  preparing  some 
grammes  of  yeast  from  substances  completely  without 
fatty  matter.  But  this  newly  formed  yeast  does  not 
contain  less  than  from  i  to  2  per  cent,  of  fatty  saponi- 
fiable  matter,  yielding  crystallized  fatty  acids.  The 
same  fact  is  observed  with  yeast  which  has  been  grown 
in  a  medium  composed  of  sugar,  water,  ammonia,  and 
phosphate.  It  is,  therefore,  from  the  elements  of  the 
sugar  that  the  fatty  matter  is  obtained. 

These  facts  confirm  the  views  of  M.  Dumas  as 
to  the  possible  formation  of  fatty  matter  from 
sugar. 

Water, — Water  is,  we  need  hardly  say,  quite  as  indis- 
pensable for  yeast,  and  the  elementary  organisms,  as  for 
higher  forms  of  life. 

According  to  Wiesner,  the  cell  of  yeast  manifests  its 
activity,  develops,  and  is  nourished  within  the  limits  of 
hydratation  comprised  between  40  and  80  per  cent,  of 
water.  Yeast  dried  with  precaution  may  regain  its 
power  when  moistened  afresh.  It  may  be  understood 
from  this  why  a  solution  of  sugar,  the  concentration  of 
which  exceeds  35  per  cent,  is  not  changed  by  ferment: 
such  a  solution  takes  from  the  cells,  by  osmose,  a  suffi- 


FUNCTIONS  OF  YEAST. 


107 


cient  quantity  of  water  to  lower  their  hydratation  below 
40  per  cent. 

Wiesner's  researches  have  also  shown  that  there  are 
two  states  of  concentration  in  which  the  phenomena  of 
the  fermentation  and  nutrition  of  yeast  attain  their 
maximum  value.  One  of  these  maxima  corresponds  to 
a  solution  of  from  2  to  4  per  cent,  of  sugar  ;  the  other 
to  a  solution  of  from  20  to  25  per  cent.  These  facts 
require  confirmation  ;  at  all  events,  there  is  at  present 
no  conclusion  to  be  drawn  from  them. 

Oxygen, — The  cells  of  the  Saccharomyces  cerevisice, 
introduced  into  a  liquid  medium  containing  oxygen  in 
solution  (pure  water,  a  saccharine  solution,  with  or 
without  nutritive  mineral  and  nitrogenous  elements), 
absorb  oxygen  with  great  rapidity,  and  develop  a 
corresponding  quantity  of  carbon  dioxide.  This  fact, 
which  constitutes  true  respiration,  comparable  to  that  of 
animals,  has  been  brought  to  light  by  M.  Pasteur.  An 
excellent  method  of  obtaining  water  completely  deoxy- 
genized,  much  more  efficaciously  than  by  boiling, 
consists  in  diffusing  through  the  water  one  or  two 
grammes  (from  15*4  to  30*8  grains)  per  litre  (176  pints 
English)  of  fresh  yeast  in  the  form  of  paste,  and 
leaving  the  liquid  undisturbed  for  from  one  to  two  hours 
at  a  temperature  of  25°  to  30°  C.  (77°  to  86°  R).  Pow- 
dered zinc  shaken  up  with  water,  containing  air  in 
solution,  gives  the  same  results. 

I  determined,  by  the  help  of  M.  Quinquand,  the 
weight  of  oxygen  absorbed  by  the  unit  of  weight  of 
yeast,  in  the  unit  of  time,  when  this  organism  is  placed 
in  water  containing  air  in  solution,  without  any  mixture 
of  nutritive  materials.   These  measurements  were  taken 


io8 


ON  FERMENTATION. 


by  an  oxymetrical  process  which  I  have  invented,  with 
the  assistance  of  M.  C.  Risler,  one  of  my  pupils.  As 
this  method  seems  to  me  hkely  to  be  serviceable  in 
researches  of  this  kind,  and  in  the  study  of  biological 
phenomena,  I  think  I  ought  to  give  the  description  of  it 
here,  even  at  the  risk  of  introducing  a  foreign  element 
into  the  examination  of  the  facts  which  now  occupy  our 
attention. 

Process  for  the  Vohtinetric  Estimation  of  Dissolved 
Oxygen. — The  process  of  measurement  of  the  quantity 
of  oxygen  dissolved  in  water,  by  means  of  a  standard 
liquid,  which  was  proposed  by  M.  Gerardin  and  myself, 
(Comp.  Rend.,  vol.  75,  p.  879),  and  which  I  have 
since  improved,  with  the  assistance  of  M.  C.  Risler, 
depends  essentially  on  the  energetic  reducing  properties 
of  sodium  hyposulphite.  This  salt*  is  obtained  with  the 
greatest  facility  by  the  action  of  a  solution  of  sodium 
bisulphite  on  zinc,  either  in  plates,  shavings,  or  powder. 
Its  formation  is  much  more  rapid  when  the  zinc  em- 
ployed is  finely  divided,  and  the  points  of  contact 
between  the  metal  and  the  solution  of  bisulphite  are 
more  numerous.  Thus,  with  powdered  zinc  employed 
in  sufficient  quantity,  and  a  very  concentrated  solution 
of  bisulphite  (marking  35""  Beaune,  and  requiring  from 
5  to  7  per  cent,  of  its  weight  of  powdered  zinc),  an 
agitation  of  from  three  to  five  minutes  is  sufficient  to 
complete  the  reaction.  This  takes  place,  with  elevation 
of  temperature,  according  to  the  following  equation : — 

3  (SO.  NaO.  HO)  +  ZN2  =  S.  NaO.  HO  +  SO  (NaO)^  +  SO  (Zn  0)2  +  H^  O. 
Sodium  bisulphite.  Sodium  hyposulphite.    Sodium  sulphite.     Zinc  sulphite.  Water. 

«  P.  Schutzenbcrgcr  on  a  new  acid  of  sulphur,  Ann.  de  Chim.  et  de  Phys., 
vol.  20,  p.  251  (4). 


FUNCTIONS  OF  YEAST. 


109 


If  the  bisulphite  used  in  the  experiment  is  concen- 
trated, there  are  deposited,  a  short  time  after  the  cooHng 
of  the  hquid,  crystals  of  the  double  zinc  and  sodium 
sulphite,  while  the  hyposulphite  formed  remains  in  solu- 
tion, still  mixed  with  the  sulphites.  The  impure  solu- 
tion of  hyposulphite  (a  mixture  of  hyposulphite  and  of 
sulphite  of  soda  and  zinc)  may  be  employed  as  it  is  for 
the  estimation  ;  but  it  will  only  keep  for  any  length  of 
time  when  protected  from  the  air,  and  in  a  very  dilute 
state. 

By  adding  to  this  liquid  a  suitable  quantity  of  milk 
of  lime,  we  precipitate  the  zinc  oxide,  and  by  filtration 
we  obtain  a  solution  very  slightly  alkaline,  endowed, 
like  the  former,  with  very  decided  reducing  power ; 
possessing  the  property  of  keeping  for  a  longer  time, 
when  not  exposed  to  the  air,  especially  in  a  state  of 
great  dilution,  the  form  under  which  it  is  always  used  in 
the  quantitative  estimation  of  oxygen.  Without  going 
at  farther  length  into  the  properties  of  sodium  hyposul- 
phite {see  the  memoir  in  the  Ann.  de  Chim.  et  de  Phys., 
before  cited),  I  ought  to  particularize  those  which  arc 
especially  utilized  in  this  process. 

I.  The  sodium  hyposulphite,  not  saturated  by  lime, 
absorbs  oxygen  with  great  rapidity,  whether  in  the 
form  of  gas,  or  in  solution  ;  its  action  is,  in  this  respect, 
similar  to  that  of  sodium  pyrogallate. 

By  fixing  the  oxygen  the  hyposulphite  becomes  acid, 
and  is  converted  unto  sodium  bisulphite. 

S.  (Na.  O)  (H  O)  +  O  =  S  O.  (Na.  O)  (H  O). 
Hyposulphite.  Bisulphite. 

When  saturated  by  lime,  it  still  acts  in  the  same 
manner  on  the  gaseous  oxygen,  but  more  slowly  ;  while 


no 


ON  FERMENTATION. 


it  absorbs  dissolved  oxygen,  instantaneously,  and  re- 
moves it  from  the  oxygenated  liquid  with  which  it  is 
mixed. 

2.  The  hyposulphite,  poured  into  a  solution  of 
ammonio-cupric  sulphate,  reduces  the  cupric  oxide  to 
the  state  of  cuprous  oxide,  destroying  the  colour  of 
the  liquid,  and  then  it  reduces  the  cuprous  oxide  in  its 
turn,  precipitating  metallic  copper  ;  the  reduction  is 
made  at  two  intervals  of  time,  and  we  are  able,  by  em- 
ploying more  or  less  hyposulphite,  to  stop  the  process 
at  the  first  stage,  shown  by  the  decolouration  of  the 
liquid. 

3.  The  hyposulphite,  whether  acid  or  neutralized, 
instantly  destroys  the  colour,  by  reduction,  of  the  solu- 
tion of  Couplers  blue  (aniline  blue),  and  of  sodium 
sulphindigotate  (indigo  carmine).  These  bleached  solu- 
tions resume  their  blue  tint  when  exposed  to  the 
air. 

4.  If  to  water  containing  oxygen  in  solution,  and 
coloured  blue  by  aniline  blue  or  indigo  carmine,  we 
add,  little  by  little,  a  dilute  solution  of  hyposulphite, 
cither  saturated  or  not  with  lime,  the  reducing  agent 
acts  at  first  upon  the  dissolved  oxygen,  and  does  not 
destroy  the  colour  until  it  has  absorbed  it. 

Thus,  by  taking  two  equal  volumes  of  water  tinted 
blue  by  cither  of  these. colouring  matters,  saturating  one 
with  oxygen  by  agitation  with  air,  and  depriving  the 
other  of  its  oxygen  by  sufficiently  prolonged  boiling, 
we  shall  find  that  the  latter  loses  it  colour  after  the 
addition  of  a  few  drops  of  the  hyposulphite,  while 
the  second  requires,  in  order  to  effect  this  result, 
a  much  greater   quantity  of  the    reducing  solution, 


FUNCTIONS  OF  YEAST. 


in 


and  one  in  proportion  to  the  quantity  of  dissolved 
oxygen. 

Here,  however,  a  very  remarkable  peculiarity  presents 
itself,  one  which  we  ought  specially  to  point  out,  be- 
cause our  ignorance  of  it  would  entail  grave  errors  in  the 
analysis.  If  we  have  some  aerated  water,  and  a  suitably 
dilute  solution  of  the  hyposulphite,  either  saturated  or 
not  with  milk  of  lime,  we  may  previously  determine  the 
oxymetric  value  of  the  hyposulphite,  that  is  to  say,  the 
volume  of  oxygen  which  is  required  to  saturate  the 
unit  of  volume  of  the  solution  ;  it  is  only  necessary  for 
this  purpose  to  prepare  a  solution  of  ammonio-cuprous 
sulphate,  containing  4*46  grammes  (68*826  grains)  of 
pure  crystallized  cupric  sulphate  to  the  litre  (176 
Eng.  pints).  Such  a  solution  having  been  brought 
exactly  to  the  bleached  state,  without  precipitation  of 
metallic  copper,  that  is  to  say,  being  brought  back  to  the 
state  of  solution  of  ammonio-cuprous -oxide,  will  have 
yielded  to  the  reducing  liquid  half  of  the  oxygen  cor- 
responding with  the  cupric  oxide  which  it  contains, 
about  I  cub.  centimetre  of  oxygen  ('o6l  cub.  in.)  for 
each  10  cub.  cent.  ('61  cub.  in.)  of  the  solution. 

It  is  sufficient,  therefore,  to  determine  with  precision 
the  volume  of  the  hyposulphite  necessary  to  decolourize 
completely  10  cubic  centimetres  of  the  cupric  liquor 
without  precipitation  of  metallic  copper ;  this  volume 
will  correspond  to  one  cubic  centimetre  of  oxygen. 

This  being  determined,  let  us  colour  with  a  little 
indigo  carmine  or  aniline  blue  (just  sufficiently  to  render 
the  tint  perceptible)  a  certain  quantity  (for  instance, 
one  or  one  half  litre)  of  our  aerated  water,  and  let  us 
pour  in  the  hyposulphite  with  a  burette  ;  the  moment 


112 


ON  FERMENTATION. 


will  come  when  the  last  drop  will  effect  the  decoloura- 
tion of  the  liquid,  which  will  rapidly  pass  from  blue  to 
yellow.  In  this  state,  the  clear  yellow  solution  is  a  very 
delicate  test  of  free  oxygen  ;  it  requires  only  the  slightest 
bubble  of  air,  of  the  size  of  a  pin*s  head  to  produce 
very  evident  blue  streaks.  We  are  therefore  induced 
to  admit  that  the  dissolved  oxygen  has  been  com- 
pletely utilized  by  the  reducing  liquid.  This  is  not, 
however,  correct.  If  we  calculate,  according  to  the 
quantity  of  hyposulphite  fixed  by  the  cupric  solution, 
and  the  volume  of  the  reducing  liquid  employed  to 
change  the  colour  of  the  blue  liquid,  the  quantity  of 
oxygen  contained  in  a  litre  of  water,  we  find,  as  nearly 
as  possible,  half  the  oxygen  really  contained  in  this 
water,  and  the  mercurial  pump  or  boiling  could  disen- 
gage from  it.  This  remarkable  result  has  been  deter- 
mined by  a  great  number  of  experiments.  What,  then, 
has  become  of  the  other  half 

M.  Risler  and  I  thought,  at  first,  that  the  products 
of  oxidation  of  the  sodium  hyposulphite  were  not  the 
same  when  the  oxidation  took  place  under  the  influence 
of  free  oxygen,  as  under  that  of  the  ammonio-cupric 
oxide  ;  however,  after  having  ascertained  that  in  both 
cases  sulphite  was  formed,  and  nothing  but  sulphite,  we 
were  obliged  to  abandon  this  interpretation  ;  we  could 
only  believe  that  the  diluted  hyposulphite,  acting,  ia  the 
cold,  on  the  dissolved  oxygen,  divides  it  into  two  equal 
parts,  one  of  which  is  fixed  in  the  reducing  liquid,  and 
the  other  unites  with  the  water,  forming  oxygenated 
water  or  some  analogous  compound.  This  second  half 
of  the  oxygen,  which  has  been,  as  it  were,  rendered 
latent,  acts  no  longer  cither  on  the  hyposulphite  or  on 


FUNCTIONS  OF  YEAST. 


the  indigo  (discoloured  carmine).  When  I  say  that  it 
no  longer  acts,  I  mean  under  the  conditions  of  the  ex- 
periment, which  may  be  considered  to  be  instantaneous, 
and  at  a  low  temperature. 

In  fact,  if  we  keep  the  bleached  liquid — provided  that 
we  have  not  used  too  little  indigo  (carmine) — for  some 
time  from  access  of  air,  especially  if  we  raise  its  tempera- 
ture to  50°  or  60°  C.  (122°  to  140''  F.),  we  see  it  become 
blue  again  instantaneously,  and  throughout  the  whole 
mass.  The  experiment  may  be  made  in  a  vessel  filled 
with  an  atmosphere  of  pure  hydrogen  to  which  is 
fixed  the  extremity  of  a  Mohr's  burette,  containing  the 
hyposulphite.  If  we  now  add  a  fresh  quantity  of  the 
reducing  fluid  until  the  second  decolouration,  the  same 
effect  will  be  produced  again,  and  until  we  have  intro- 
duced a  volume  of  hyposulphite  nearly  equal  to  that 
employed  in  order  to  attain  the  first  term  of  decolour- 
ation. 

These  experiments  are  delicate.  To  make  them 
succeed  they  must  be  completely  protected  from  the 
access  of  atmospheric  oxygen ;  hyposulphite  which  has 
been  neutralized  by  lime,  should  be  employed,  and  there 
should  be  a  sufficient  quantity  of  indigo.  (For  further 
details  on  this  subject,  see  the  Bulletin  de  la  Societe 
Chim.  de  Paris,  vol.  20,  p.  145,  1873.)  They  prove  that 
the  first  action  of  the  acid  hydrosulphite  (which  may  be 
considered  instantaneous)  on  the  aerated  water  coloured 
by  indigo,  only  removes  half  the  oxygen.  The  other 
half  acts  much  more  slowly  on  the  reduced  indigo,  and, 
by  its  intervention^  on  the  hyposulphite  in  excess.  This 
action  does  not  manifest  itself  at  all  if  the  solution  is 
even  slightly  acid.  In  this  case,  the  latent  oxygen 
6 


114 


ON  FERMENTATION 


may  remain  almost  indefinitely  in  the  presence  of  a 
great  excess  of  hyposulphite,  or  of  the  reduced  solution 
of  indigo  carmine,  without  being  fixed  by  it. 

By  employing  water  coloured  blue,  to  which  has  been 
added  a  little  oxygenated  water  (H^  O^),  we  produce 
with  the  hyposulphite  alternate  decolourations,  followed 
by  spontaneous  recolourations,  of  the  whole  mass,  which 
resemble,  so  as  to  be  indistinguishable  from  them,  those 
which  take  place  in  the  experiments  already  described. 
This  similarity,  added  to  the  want  of  any  other  plausible 
explanation,  makes  me  think  that  the  latent  oxygen  is 
really  found  in  the  liquid  under  the  form  of  oxygenated 
water. 

If  we  operate  on  a  liquor  rather  acid  than  neutral,  or 
on  a  neutral  liquor  employing  only  hyposulphite  not 
saturated  with  lime,  which  becomes  acid  as  it  oxidizes  ; 
finally,  by  taking  note  of  the  previous  observation  in 
our  calculation,  that  is,  multiplying  the  quantity  of 
oxygen  found  by  2,  we  arrive  at  very  close  and  satis- 
factory results. 

I  will  first  describe  a  rough  method,  susceptible  of 
being  used  anywhere,  on  the  banks  of  a  river,  or  in  the 
country,  but  which  can  furnish  only  approximate  indica- 
tions, by  giving  the  amount  of  oxygen  within  a  quarter 
of  a  cubic  centimetre  per  litre  (*oi5  cub.  in.  per  1*76 
pint). 

Some  acid  hyposulphite  may  be  prepared,  instan- 
taneously, by  agitating  with  zinc  powder  a  diluted 
solution  of  sodium  bisulphite,  prepared  with  super- 
saturating sodium  carbonate  by  a  current  of  sulphurous 
acid  (the  bisulphite  at  35°  Beaume  is  a  commercial  pro- 
duct, and  may  be  used).    This  bisulphite  at  35°  Beaumd 


FUNCTIONS  OF  YEAST. 


is  previously  diluted  with  four  times  its  weight  of  water 
and  for  100  grammes  of  the  diluted  solution  we  em- 
ploy 2  grammes  of  zinc  grey  (powdered  zinc).  The 
mixture  and  the  agitation  are  to  be  made  in  a  vessel 
nearly  filled  with  the  liquid.  After  five  minutes,  the 
solution  must  be  filtered,  and  suitably  diluted  with  water, 
so  that  in  a  preliminary  trial,  one  litre  of  water  agitated 
with  air  (saturated  with  oxygen  under  the  pressure  of 
3  of  an  atmosphere  at  the  ordinary  temperature)  and 
tinted  blue  by  some  drops  of  a  solution  of  aniline  blue,  or 
indigo  carmine,  may  be  decoloured  by  about  25  or  35 
cubic  centimetres  (1*525  to  2135  cubic  in.)  of  the  solu- 
tion of  hyposulphite. 

The  analysis  requires  nothing  but  a  vessel  with  a 
large  mouth,  (a  wide-mouthed  bottle),  holding  about 
I  y2  litre,  a  stirrer  which  will  allow  us  to  mix  together 
the  different  layers  of  liquid  without  disturbing  the 
surface  too  much,  one  of  Mohr's  burettes,  furnished 
with  a  narrow  tube  at  one  end,  fixed  to  the  india- 
rubber  tube  of  the  pinch-cock,  and  arranged  so  as  to 
be  held  mid-way  in  the  water ;  also  a  glass  bottle  or 
jar  holding  a  little  more  than  two  litres,  graduated  so 
as  to  indicate  i  litre.  A  litre  (about  pints) 
of  the  water  to  be  tested,  is  introduced  into  the  wide- 
mouthed  bottle,  tinted  with  aniline  blue  or  indigo 
carmine  ;  then,  the  burette  being  filled  with  hyposul- 
phite, and  its  lower  end  previously  filled  with  the 
liquid  plunging  midway  into  the  water  in  the  wide- 
mouthed  bottle,  we  allow  the  reducing  liquid  to  flow 
in  slowly,  agitating  the  contents  with  the  stirrer  up 
and  down,  so  as  not  to  disturb  the  surface  too  much ; 
the  experiment  is  stopped  at  the  moment  that  the 


ii6 


ON  FERMENTATION. 


decolouration  takes  place,  and  the  volume  employed  is 
read  off. 

Immediately  after,  we  proceed  to  the  estimation  of 
the  hyposulphite  exactly  in  the  same  manner,  by 
employing  I  litre  of  the  same  kind  of  water  which 
served  for  the  first  experiment,  but  after  having  pre- 
viously agitated  it  for  some  minutes  with  air,  in  the 
large  bottle,  and  taken  its  temperature.  Under  these 
conditions,  whether  the  original  water  be  above  or 
below  the  limit  of  saturation  for  oxygen  ;  we  always 
succeed  quickly  in  having  water  saturated  with  oxygen 
at  the  pressure  of  ^  of  an  atmosphere  (the  pressure  of 
oxygen  in  the  air),  and  at  the  temperature  which  has 
been  read  off.  Tables  of  solubility,  notably  those  of 
Bunsen  (Methodes  Gazometriques,  Traduction  Frangaisc 
de  T.  Schneider)  give  the  amount  of  oxygen. 

Thus,  in  two  experiments  made  under  conditions 
identically  similar,  we  have  the  volume  of  the  reducing 
fluid  required  by  the  water  whose  oxygen  is  unknown, 
and  that  required  by  the  water  whose  oxygen  is  known. 
A  simple  proportion  will  give  the  value  of  x  in  the  pro- 
blem. This  process  of  estimating  the  hyposulphite,  on 
account  of  its  simplicity  and  certainty,  is  preferable  to 
the  employment  of  an  ammoniacal  solution  of  copper, 
which  M.  Gerardin  and  I  had  proposed  ;  it  was  sug- 
gested by  M.  Raulin,  assistant  director  of  the  labora- 
tory of  M.  Pasteur.  As  the  operation  is  performed  with 
contact  of  air,  it  is  necessary  to  make  the  measure- 
ments as  quickly  as  possible,  and  to  operate  on  a  large 
quantity  of  water  (a  litre),  in  order  to  counteract  as  far 
as  possible  the  influence  of  the  oxygen  of  the  air. 
Besides,  the  method  of  analysis  described  above  has 


FUNCTIONS  OF  YEAST. 


117 


the  effect  of  neutralizing  almost  entirely  this  cause  of 
error ;  the  two  operations  being  made  under  the  same 
conditions,  the  error  can  only  proceed  from  a  slight 
difference  between  the  conditions  of  the  two  experi- 
ments, such  as  their  duration,  or  the  greater  or  less 
agitation  of  the  water. 

I  have  succeeded,  by  the  assistance  of  M.  Risler,  in 
applying  a  similar  method  of  quantitative  analysis  to 
much  smaller  quantities  of  water,  or  oxygenated  liquid  ; 
and  by  modifying  the  process  of  the  operation,  I  have 
been  able  to  ascertain  by  the  reducing  liquid,  not  the 
halfy  but  the  whole  of  the  dissolved  oxygen ;  this 
method  is  much  preferable  and  more  certafn.  In  order 
to  attain  this  double  result  it  is  only  requisite  ;  ist.  To 
make  the  analysis  in  a  liquid  completely  protected  from 
access  of  the  oxygen  in  the  air,  by  an  atmosphere  of 
pure  hydrogen :  2nd.  To  introduce  the  aerated  water  that 
is  to  be  tested,  (a  known  volume,  from  40  to  100  cubic 
centimetres  (2*44  to  6*1  cub.  in.)  into  a  tepid,  40°  or  50*^ 
C.  (104''  to  122°  F.)  neutral,  or  very  slightly  alkahne, 
but  never  acid,  piedium,  formed  by  a  solution  of  in- 
digo carmine,  just  decoloured  by  hyposulphite  which 
has  been  previously  neutralized  or  rendered  slightly 
alkaline  by  milk  of  lime.  This  yellow  medium  turns 
blue  under  the  influence  of  dissolved  oxygen ;  a 
quantity  of  blue  indigo  is  re-formed,  proportional  to 
the  amount  of  oxygen  dissolved.  If  the  preceding 
conditions  of  temperature  and  neutralization  have  been 
carefully  observed,  all  the  dissolved  oxygen  is  utilized 
in  oxygenating  the  reduced  indigo,  and  there  only 
remains  to  be  estimated,  by  means  of  the  hyposulphite, 
the  volume  of  this  reducing  agent  necessary  to  de- 


Il8  ON  FERMENTATION. 

colour  the  blue  liquid.  The  same  experiment  is  re- 
peated immediately  after,  with  the  same  volume  of 
water,  agitated  at  a  known  temperature,  and  calculation 
will  give,  as  before,  the  volume  of  oxygen  sought.  If 
the  liquid  is  acid,  or  becomes  so  in  the  process  of 
analysis,  the  conditions  leading  to  the  formation  of 
oxygenated  water  are  at  once  present,  and  the  results 
are  deceptive,  and  always  too  low,  approaching  more 
or  less  nearly  to  the  half  of  the  whole  amount  of 
oxygen. 

If,  then,  we  operate  on  acid  liquids^,  we  ought  to 
add  previously  to  the  yellow  indigotic  liquid  a  sufficient 
quantity  of  a  diluted  solution  of  ammonia  to  correct 
this  disturbing  condition.  The  principles  of  the  ex- 
periment being  known,  I  will  enter  into  some  details 
concerning  the  apparatus,  and  the  preparation  of  the 
reagents. 

I.  Sodmm  Hyposidphiie, — We  prepare  the  acid  hy- 
posulphite in  the  manner  described  above.  This  is 
neutralized  by  milk  of  lime.  For  lOO  grammes  of 
concentrated  bisulphite  at  35°  Beaume,  employed  for 
the  preparation  of  the  acid  hyposulphite,  we  shall 
make  use,  for  this  purpose,  of  35  grammes  of  milk  of 
lime,  prepared  from  200  grammes  of  lime  previously 
slaked,  to  i  litre  of  water.  The  saturation  is  effected 
with  the  acid  hyposulphite,  diluted,  as  I  have  before 
said,  with  four  times  its  weight  of  water,  but  the 
quantity  of  milk  of  lime  is  calculated  according  to 
the  weight  of  concentrated  bisulphite  that  is  employed. 
It  is  shaken  up,  left  to  settle,  decanted,  and  then 
filtered  ;  the  liquid  is  kept  in  full  and  well-stoppered 
bottles. 


FUNCTIONS  OF  YEAST. 


119 


When  used,  it  is  necessary  to  dilute  this  liquid  with 
distilled  water,  until  50  cubic  centimetres  (3*05  cub.  in.), 
of  water  saturated  with  oxygen  require  from  4  to  5 
cubic  centimetres  ('244  to  '305  in.),  of  the  reducing 
agent. 

The  solution  thus  prepared  may  be  kept  almost  in- 
definitely at  the  required  strength,  if  we  take  the  fol- 
lowing precautions.  It  is  to  be  placed  in  a  bottle  con- 
taining about  I  litre  (i^  pints)  filled  to  the  neck,  closed 
by  a  good  india-rubber  cork,  perforated  with  two  holes  ; 
in  one  of  them  is  fixed  a  tube  bent  at  right  angles, 
whose  extremity  dips  to  the  bottom  of  the  bottle,  and 
whose  other  end  has  an  india-rubber  tube,  furnished 
with  a  Mohr's  pinch  cock  ;  in  the  free  end  of  the  india- 
rubber  tube,  is  fixed  a  piece  of  glass  tube ;  in  the 
second  hole  is  fixed  another  tube  bent  at  right  angles, 
but  which  only  goes  about  i  or  2  centimetres  ('39  or 
78  in.)  past  the  cork.  This  tube  is  kept  in  permanent 
communication  by  means  of  a  sufficiently  long  india- 
rubber  tube,  with  a  gas-jet  kept  turned  on.  It  is  proper 
to  interpose  between  the  gas-burner  and  the  bottle  a 
glass  vessel  like  those  employed  by  chemists  for  drying 
gases  ;  this  vessel  is  filled  with  pumice  stone,  saturated 
with  a  concentrated  solution  of  sodium  pyrogallate. 
By  this  means,  we  get  rid  of  the  oxygen  which  common 
gas  always  contains  (from  i  to  2  per  cent.)  The 
burettes  are  filled  with  hyposulphite  by  aspiration,  from 
below  upwards.  For  this  purpose,  the  india-rubber 
pinch-cock  tube,  which,  in  order  to  serve  another  pur- 
pose, ought  to  be  sufficiently  long,  is  placed  in  commu- 
nication with  the  tube  which  goes  to  the  bottom  of  the 
hyposulphite,  and  we  draw  in  the  liquid  by  the  mouth, 


I20 


ON  FERMENTATION. 


by  means  of  an  india-rubber  tube,  fixed  by  a  cork  and 
a  bent  glass  tube  to  the  upper  part  of  the  Mohr  s 
burette.  By  this  means,  we  avoid  the  agitation  of  the 
liquid  in  contact  with  the  air.  With  these  precautions, 
the  strength  of  the  test  does  not  alter  ;  it  is,  however, 
prudent  not  to  trust  to  this,  but  to  ascertain  it  at  every 
fresh  experiment,  which  is  very  easily  done. 

Indigo. — We  prepare  beforehand  ten  litres  of  the 
solution  of  indigo  carmine,  by  dissolving  in  this  quan- 
tity nearly  200  grammes  (6*42  oz.  troy),  of  indigo 
carmine  in  the  form  of  paste  (sodium  sulphindigotate). 
The  liquid  should  be  kept  in  blue  or  black  glass  bottles, 
sheltered  from  the  light. 

The  estimation  is  made  in  a  three-necked  bottle,  of 
I  or  ij^  litres  in  capacity  (Fig.  19).  One  of  the  three 
lateral  necks  receives,  through  an  india-rubber  cork,  the 
tube,  Hy.,  bent  at  right  angles  for  the  admission  of 
hydrogen.  It  is  well  that  this  tube  should  slide  in  its 
cork  without  too  much  friction,  in  order  that  it  may 
be  raised  or  depressed  at  will,  without  allowing  air  to 
enter. 

The  other  lateral  neck  has  an  india-rubber  cork, 
pierced  with  two  holes.  In  one  of  these  is  fixed  the 
end  of  a  small  funnel  with  a  glass  stop-cock,  E,  a 
bromide  funnel  ;  this  end  should  be  sufficiently  long  to 
reach  the  bottom  of  the  bottle. 

In  the  other  hole  is  fixed  the  tube  which  carries  off 
the  hydrogen  ;  this  tube,  being  twice  bent,  has  its  free 
extremity  plunged  into  the  mouth  of  a  test-tube,  T, 
which  is  fixed  there  by  means  of  a  cork,  also  pierced 
with  two  holes  ;  the  tube  contains  water,  and  the 
hydrogen  in  excess,  after  having  passed  through  this 


FUNCTIONS  OF  YEAST. 


121 


water,  escapes  by  a  tube  bent  at  right  angles,  fixed  in 
the  second  hole  of  fhe  cork. 

The  middle  neck  of  the  bottle  holds  as  a  fixture  a 
stopper  of  cork  or  india-rubber,  hermetically  cemented 
in,  and  pierced  with  two  orifices,  in  w^hich  are  also  per- 
manently fixed  the  two  pointed  extremities  of  two 
Mohr  burettes,  H  I,  held  above  the  bottle  by  a  special 


Fig.  19.—  Apparatus  for  the  measurement  of  oxygen  dissolved  in  water. 

support,  one  by  the  side  of  the  other.  The  india- 
rubber  pinch-cock  tubes  at  the  lower  end  of  these 
two  burettes  ought  to  be  long  enough  to  allow  the 
bottle  to  be  moved  without  the  burettes  being  shaken  ; 


122 


ON  FERMENTATION. 


they  are  fixed  to  the  burettes,  and  ought,  on  the  con- 
trary, to  be  able  to  be  detached,  at  will,  from  the  thin 
glass  ends  which  pass  through  the  middle  cork  of  the 
bottle.  It  is  well  to  have  at  hand  a  third  burette,  either 
independent,  or  fixed  to  the  same  support  by  the  side 
of  the  others.  It  is  intended  as  a  reserve  for  the  hypo- 
sulphite of  the  principal  burette,  H.  One  of  the  first 
two  burettes,  I,  contains  indigo  carmine,  the  two  others 
are  filled  with  sodium  hyposulphite. 

This  having  been  arranged,  when  we  wish  to  measure 
the  oxygen  dissolved  in  any  particular  specimen  of 
water,  we  introduce  into  the  large  bottle — 

(1)  About  50  cubic  centimetres  (3*05  cub.  in.)  of 
solution  of  indigo. 

(2)  About  250  cubic  centimetres  (15*25  cub.  in.)  of  or- 
dinary warm  water,  at  40°  to  50°  C.  (104°  to  122°  F.).  We 
adapt  to  the  fixed  sockets  the  burette,  I,  with  the  indigo, 
and  the  independent  burette  of  hyposulphite  ;  we  charge 
the  two  sockets  with  the  contents  of  the  burettes,  and 
allow  the  hydrogen  to  pass  rather  quickly.  (The  appa- 
ratus producing  the  hydrogen  may  be  of  any  kind, 
provided  that  we  are  able  to  regulate  the  disengagement 
of  gas  by  means  of  a  single  stop-cock  ;  the  hydrogen 
is  purified  by  water,  and  passes  through  a  column  of 
caustic  potash  in  plates.)  When  we  suppose  that  almost 
the  whole  of  the  air  has  been  swept  away  by  the  current 
of  hydrogen,  we  allow  the  hyposulphite  to  flow  in 
slowly,  holding  the  bottle  in  the  hand,  and  giving  the 
liquid  a  gyratory  motion  sufficient  to  mix  it,  until  it  has 
only  a  slight  greenish  tint,  or  reddish  if  it  is  alkaline  ; 
then,  without  interrupting  the  current  of  hydrogen  gas, 
we  detach  the  independent  burette,  and  fix  on  the  hypo- 


FUNCTIONS  OF  YEAST. 


123 


sulphite  burette,  H,  intended  for  the  analysis.  In  doing 
this,  the  test  liquid  contained  in  the  socket  escapes,  and 
often  completes  the  decolouration  of  the  water  by  turn- 
ing it  yellow.  We  charge  the  socket  again  by  slightly 
opening  the  pinch-cock,  then  by  adding  first  a  little 
carmine,  then  a  little  hyposulphite,  we  bring  the  liquor 
to  a  bright  yellow  colour,  which  verges  towards  green 
or  red  by  the  addition  of  a  single  drop  of  carmine. 
We  notice  that  the  yellow  liquid  does  not  turn  blue  at 
the  surface,  which  proves  that  the  atmosphere  of  the 
bottle  is  thoroughly  deprived  of  oxygen.  Then  all  is 
ready  for  the  analysis  if  we  have  taken  care,  at  first, 
to  fill  the  lower  end  of  the  funnel  with  the  same  water 
as  that  whose  contained  oxygen  we  wish  to  measure. 
This  estimation  is  made  by  proceeding  in  the  following 
manner,  and  in  the  order  indicated : — 

(i)  Slacken  the  current  of  hydrogen,  without  stopping 
it ;  (2)  Raise  the  supply  tube,  so  that  the  gas  should 
not  bubble  up ;  (3)  Read  off  the  graduation  of  the 
hyposulphite  burette,  at  which  the  surface  of  the  liquid 
stands  ;  (4)  Introduce  into  the  funnel  50  or  100  cubic 
centimetres  of  the  water  to  be  tested,  and  allow  this 
water  to  run  at  once  into  the  bottle,  keeping  the  lower  end 
full  from  the  stop-cock  ;  shake  the  bottle  ;  the  standard 
yellow  liquid  grows  blue  if  the  water  is  aerated,  and 
does  not  change  its  colour  if  it  contains  no  oxygen. 
We  now  need  only  allow  the  hyposulphite  to  run  in, 
drop  by  drop,  shaking  it,  and  noticing  the  exact  limit  of 
decolouration  which  takes  place,  true  to  a  single  drop 
(2V  of  a  cubic  centimetre)  if  the  liquid  is  without  colour. 

Immediately  after  having  noted  the  point  at  which 
this  takes  place,  without  changing  or  disturbing  any- 


124 


ON  FERMENTATION. 


thing,  fill  the  body  of  the  funnel  with  water  saturated 
with  oxygen,  at  a  pressure  of  }  of  an  atmosphere,  and 
at  a  known  temperature ;  correct  with  some  drops  of 
hyposulphite  the  slightly  blue  tint  developed  by  this 
operation,  and  proceed  to  estimate  the  value  with  50 
or  100  cubic  centimetres  of  saturated  water.  A  rule 
of  three  sum  gives  the  quantity  of  oxygen  in  the  first 
portion  of  water. 

Instead  of  testing  the  hyposulphite  with  saturated 
water  at  a  known  temperature,  we  may  estimate  it  by 
allowing  a  known  volume  (25  cubic  centimetres)  of  solu- 
tion of  indigo  carmine  to  flow  into  the  bottle  after  the 
first  trial,  and  ascertaining  the  volume  of  the  reducing 
agent  necessary  to  bring  back  the  decolouration. 

On  the  other  hand,  we  have  determined,  once  for  all, 
the  volumetric  ratio  between  the  indigo  employed,  and 
a  solution  of  ammonio-cupric  sulphate,  containing  4*46 
grammes  of  pure  crystallized  cupric  sulphate  to  the  litre ; 
this  amounts  to  10  cubic  centimetres  =  i  cubic  centi- 
metre of  oxygen  at  the  moment  of  complete  decoloura- 
tion. It  is  sufficient  for  this  purpose  to  ascertain  the 
volumes  of  the  same  solution  of  hyposulphite  necessary 
to  decolour  equal  volumes  of  indigo  carmine  and  of  the 
cupric  liquid,  which  allows  us  to  calculate  the  ratios  of 
volumetric  equivalence  of  these  two  liquids,  and  con- 
sequently the  volume  of  oxygen  corresponding  to  I 
cubic  centimetre  of  the  indigo  solution. 

This  experiment  ought  to  be  made  with  great  care, 
for  on  its  accuracy  will  depend  that  of  the  absolute 
values  of  all  the  subsequent  analyses. 

We  determine  the  ratio  between  the  hyposulphite 
and  the  indigo  under  the  same  conditions  as  those  of 


FUNCTIONS  OF  YEAST. 


the  preceding  estimation,  since  these  are  the  conditions 
to  which  we  shall  always  come  back  in  future  experi- 
ments. 

The  ratio  between  the  same  hyposulphite  and  the 
cupric  solution  is  determined  by  operating  in  an  atmos- 
phere of  pure  hydrogen.  The  cupric  solution  (15  to  20 
cubic  centimetres)  is  placed  in  a  small  bottle  with  three 
necks,  one  for  the  admission  the  other  for  the  exit  of 
the  gas,  this  latter  being  furnished  with  a  small  stirring 
apparatus,  similar  to  that  on  the  large  bottle  ;  on  the 
middle  neck  of  the  bottle  is  fixed,  by  an  india-rubber 
tube,  the  socket  of  the  hyposulphite  burette,  lengthened 
and  drawn  out  to  a  fine  point,  and  charged  beforehand. 
The  reducing  agent  is  allowed  to  run  in  as  soon  as  the 
air  is  expelled,  and  this  is  continued  till  complete  de- 
colouration takes  place  ;  this  point  is  somewhat  difficult 
to  ascertain. 

Example, — By  operating  as  above,  we  have  found 
that— 

(1)  4*6  cubic  centimetres  of  hyposulphite  were  equiva- 
lent to  50  cubic  centimetres  of  indigo. 

(2)  15*1  cubic  centimetres  of  hyposulphite  were 
equivalent  to  25  cubic  centimetres  of  ammoniacal  solu- 
tion of  copper;  10  cubic  centimetres  of  decoloured 
cupric  solution  z=  i  cubic  centimetre  of  oxygen  (at  o"" 
C.  (32''  R),  and  760  millimetres  of  pressure).  We  easily 
find  that  I  cubic  centimetre  of  indigo,  corresponds  to 
0'0i52  cubic  centimetres  of  oxygen. 

On  the  other  hand  ; 

For  100  cubic  centimetres  of  water  to  be  tested,  we 
have  employed  57  cubic  centimetres  of  some  hypo- 
sulphite. 


126 


ON  FERMENTATION. 


20  cubic  centimetres  of  indigo  require  2*9  cubic  centi- 
metres of  this  hyposulphite. 
We  make  the  proportion  : — 

2*9  :  20  :  :  57  :  x=^-^^^  =  39*3  cubic  centimetres. 

The  oxygen  of  100  cubic  centimetres  of  water  corre- 
sponds to  39*3  cubic  centimetres  of  indigo.  Nothing 
remains  but  to  multiply  39*3  by  0*0152  =:  0*59736,  to 
find  that  one  litre  of  water  contained  5*97  cubic  centi- 
metres of  dissolved  oxygen,  measured  at  o""  C.  (32*'  F.) 
and  at  760  millimetres  pressure. 

N.B. — It  is  important,  when  we  make  a  series  of 
analyses,  never  to  use  the  last  cubic  centimetres  of  the 
burette  of  hyposulphite,  which,  having  remained  in  con- 
tact with  the  air,  have  lost  their  reducing  power.  If  we 
leave  more  than  half  an  hour's  interval  between  the  trials, 
it  would  be  better  to  renew  entirely  the  contents  of  the 
burette. 

A  complete  experiment,  including  the  preparation  of 
the  reduced  medium,  does  not  require  more  than  ten 
minutes,  and  each  of  the  succeeding  measurements  is 
made  in  one  or  two  minutes.  Two  trials  of  the  same 
kind,  repeated,  never  differ  more  than  -i-q  of  a  cubic 
centimetre. 

This  process  of  analysis  allowing  us  to  estimate  the 
oxygen  dissolved  in  50  cubic  centimetres  of  water  (3*05 
cub.  in.)  with  an  approximation  of  0*005  cubic  centi- 
metre, and  consequently  of  O'l  cubic  centimetre  per 
litre,  we  have  been  able  to  utilize  it  for  the  purpose  of 
studying  the  respiratory  phenomena  of  yeast,  and  of 
measuring  their  intensity  under  different  conditions  of 
temperature.    The  rapidity  of  the  estimates,  which  only 


FUNCTIONS  OF  YEAST. 


127 


require  three  or  four  minutes,  gave  us  the  opportunity 
of  multiplying  the  experiments,  and  establishing  the 
results  which  are  given  below  by  a  series  of  analyses, 
the  number  of  which  w^as  not  obliged  to  be  restricted. 
The  experiments  to  which  I  shall  here  allude  apply  only 
to  the  case  of  fresh  yeast  in  the  form  of  paste,  containing 
from  29  to  30  per  cent,  of  solid  matter,  diffused  in  pure 
aerated  water,  without  the  addition  of  any  nutritive 
element.  We  shall  presently  see  what  takes  place 
when  the  yeast  is  placed  under  conditions  more  favour- 
able to  its  developement. 

The  method  consists  in  leaving  a  known  weight  of 
yeast  for  a  known  length  of  time  in  contact  with  a 
known  weight  of  water,  under  the  conditions  of  temper- 
ature which  we  may  wish  to  study.  The  oxymetrical 
degrees  of  the  water  are  measured  at  the  commence- 
ment and  the  end  of  the  experiment.  Their  difference 
gives  the  oxygen  absorbed. 

The  yeast  of  beer  only  shows  the  phenomenon  of  ab- 
sorption of  oxygen,  with  production  of  carbon  dioxide. 
All  things  else  being  equal,  the  respiratory  intensity 
is  the  same  in  the  dark,  in  diffused,  and  in  full  light ; 
it  is  proportionate  to  the  weight  of  yeast  employed. 

The  original  amount  of  dissolved  oxygen  does  not 
sensibly  influence  the  results,  except  when  it  is  below 
I  cubic  centimetre  per  litre.  We  find,  in  this  case,  a 
feeble  diminution  in  the  rapidity  of  the  absorption, 
which  continues  till  complete  deoxygenation  of  the 
water. 

Below  10''  C.  (50°  F.),  the  absorbing  power  of  the 
yeast  for  oxygen  is  almost  nil ;  it  increases  slowly  to 
iS""  C.  (about  65°  F.)  ;  from  this  point  the  increase  is 


128 


ON  FERMENTATION. 


rapid  till  about  35°  C.  (95°  R),  a  temperature  at  which 

 the  respiratory  intensity  attains  its  maximum,  which  is 

sustained  sensibly  till  50°  C.  (122°  F.).  At  Gd"  C.  (140° 
F.),  the  absorbing  power  is  annulled  and  destroyed. 

A  specimen  of  yeast,  sensibly  fresh,  containing  26 
per  cent,  of  solid  matter,  absorbed,  per  gramme  and  per 
hour,  at  C.  (about  49''  F.),  0*14  cubic  centimetre  ; 
at  11^  (52"  F.),  0*42  cubic  centimetre  ;  at  22^  C.  (about 
72^  F.),  1*2  cubic  centimetre;  at  33°  C.  (about  93""  F.), 
2*1  cubic  centimetre;  at  40°  C.  (104°  F.),  2*06  cubic 
centimetres  ;  at  50""  C.  (122''  F.),  2*4  cubic  centimetres  ;  at 
60"  C.  (140°  F.),  none. 

Another  specimen  of  yeast,  very  fresh,  and  of  very 
good  appearance,  containing  30  per  cent,  of  solid  matter, 
absorbed,  per  gramme  and  per  hour,  at  24^  C.  (about 
76^  F.),  2*2  cubic  centimetres  of  oxygen ;  at  36®  C. 
(about  97**  F.)  107  cubic  centimetres. 

The  increase  of  absorbing  power  between  24**  C.  and 
36^  C.  was  therefore  more  considerable  than  with  the 
first  yeast ;  this  power  is  doubled  in  the  one  case,  and 
quintupled  in  the  other. 

The  values  of  the  quantities  of  the  oxygen  absorbed, 
as  well  as  the  magnitude  of  the  variations  with  relation 
to  the  temperature,  are  by  no  means  absolute ;  they 
depend  on  a  particular  factor,  inherent  in  the  yeast, 
which  we  may  call  the  factor  of  vitality;  but  what- 
ever this  factor  may  be,  the  direction  of  the  variations 
is  always  the  same,  and  susceptible  of  being  repre- 
sented by  a  curve  starting  from  the  line  of  abscissae,  or 
line  of  temperature,  at  about  (f  or  io«  C.  (about  40''  or 
50''  F.),  rising  slowly  up  to  18"  C.  (about  65^  F.),  thence 
rapidly  reaching,  at  about  35^  C.  (95°  F.),  a  maximum 


FUNCTIONS  OF  YEAST. 


129 


height,  which  it  retains  till  nearly  60°  C.  (140''  F.),  when 
it  returns  suddenly  towards  the  line  of  abscissae. 

Yeast  can  not  only  utilize  and  cause  to  disappear  the 
oxygen  physically  dissolved  in  water,  but  also  oxygen 
combined  with  hsemaglobin,  which,  as  we  know,  can  be 
eliminated  by  a  diminution  of  pressure. 

Thus,  when  we  diffuse  fresh  yeast,  whether  washed  or 
not,  in  arterial  red  blood,  or  in  a  solution  of  haemaglobin 
saturated  with  oxygen,  we  see  the  tint  change  rapidly 
from  red  to  dark  blue  or  black.  A  simple  agitation 
of  the  blood  with  air  is  sufficient  to  restore  its  ruddy 
colour ;  then  the  phenomena  of  deoxygenation  recom- 
mence; the  same  experiment  may  thus  be  repeated  a  great 
number  of  times,  especially  with  fresh  and  washed  yeast. 

Although,  in  this  case,  the  yeast  is  in  contact  with  a 
medium  infinitely  more  rich  in  oxygen  than  is  aerated 
water  (containing  from  200  to  230  cubic  centimetres  of 
oxygen  to  the  litre,  instead  of  from  6  to  7  cubic  centi- 
metres), the  rapidity  of  the  absorption  is  not  increased, 
if  the  conditions  of  temperature  are  the  same. 

One  gramme  of  yeast  absorbs  as  much  oxygen  in  an 
hour,  at  the  same  temperature,  whether  it  be  mixed  with 
water  containing  5  or  7  cubic  centimetres  of  oxygen  per 
litre,  or  in  arterial  blood  containing  200  cubic  centimetres 
of  oxygen. 

In  the  experiment  with  blood,  we  might  fear  a  direct 
influence  of  the  yeast,  or  of  its  soluble  materials,  on  the 
colouring  matter  of  blood ;  this  influence  is,  in  fact, 
produced,  especially  with  solutions  of  haemaglobin  ;  it 
shows  itself  by  the  transformation  of  this  primordial 
colouring  matter  into  haematin  ;  but  it  only  makes  its 
appearance  after  some  hours. 


130 


ON  FERMENTATION. 


The  behaviour  of  the  yeast  with  reference  to  blood 
may  be  explained  in  the  following  manner  :  The  cells 
of  SaccJiaromyces  diffused  in  the  liquid  breathe  at  the 
expense  of  the  oxygen  physically  dissolved  in  the  plasm 
or  serum  in  the  midst  of  which  swim  the  red  globules 
of  blood.  In  proportion  as  the  plasmic  liquid  grows 
less  rich  in  oxygen,  a  portion  of  this  body,  feebly  com- 
bined with  hsemaglobin,  is  separated,  and  enters  into 
physical  dissolution,  by  a  dissociation  comparable  to 
that  presented  by  potassium  bicarbonate  in  a  vacuum  ; 
the  process  continues  till  there  is  a  complete  disappear- 
ance of  the  oxygen  dissolved  in  the  serum,  and  of  that 
which  is  fixed  in  the  haemaglobin.  If  this  explanation 
is  correct,  the  experiment  ought  to  succeed,  even  when 
the  blood  is  separated  from  the  yeast  diffused  in  water 
or  serum  by  means  of  a  membrane  permeable  to 
gas  and  to  liquids,  but  capable  of  preventing  all  direct 
contact  between  the  yeast-cells  and  the  red  globules. 
This  is,  in  fact,  what  takes  place.  I  have  thus  been 
able,  by  arranging  a  suitable  apparatus,  to  imitate  arti- 
ficially that  which  takes  place  in  the  organs  and  tissues 
of  animals,  when  the  red  and  oxygenated  arterial  blood 
traverses  the  network  of  capillary  vessels,  and  passes 
out  into  the  veins  under  the  form  of  black  and  partially 
deoxygenated  blood. 

For  this  purpose,  it  is  only  necessary  to  cause  red 
blood  to  circulate  slowly  through  a  sufficiently  long 
system  of  hollow  tubes,  the  walls  of  which  are  formed 
of  thin  gold-beater's  skin,  which  is  immersed  in  a  mix- 
ture of  yeast  diffused  in  fresh  scrum,  without  globules, 
kept  at  35°  C.  (95°  F.). 

We  see  the  red  blood  pass  out  black  and  venous  at 


FUNCTIONS  OF  YEAST. 


the  other  extremity.  A  confirmatory  experiment,  made 
at  the  same  time,  with  a  system  of  tubes  precisely 
similar,  but  immersed  in  serum  without  yeast,  proves 
that  yeast  is  indispensable  for  thus  rapidly  effecting  the 
deoxidation  of  the  blood.  This  experiment  is  the 
exact  representation  of  what  takes  place  in  the  animal 
organism,  with  the  exception  of  the  perfect  method 
employed  by  nature  to  multiply  contacts  and  surfaces. 

In  the  latter  case,  the  cellular  and  histological  ele- 
ments of  the  tissues  play  the  part  of  the  yeast ;  they 
absorb  the  oxygen  dissolved  in  the  plasmic  liquids  which 
bathe  them,  and  constantly  tend  to  bring  down  to  zero 
their  oxymetric  condition.  The  oxgen,  but  feebly  fixed 
in  the  hsemaglobin,  re-establishes  the  equilibrium  by  a 
series  of  gaseous  diffusions  from  the  red  globules  to  the 
plasm  of  the  blood,  and  from  the  plasm  of  the  blood  to 
that  of  the  organs.  These  continual  diffusions  are  the 
inevitable  consequence  of  the  disturbance  of  equilibrium 
produced  by  the  aeration  of  the  organic  cells,  or  of 
the  cells  of  yeast  in  the  experiment  just  described. 

All  these  facts,  then,  prove  distinctly  that  yeast 
breathes  when  placed  in  contact  with  dissolved  oxygen 
The  measure  of  the  respiratory  power,  under  the  most 
favourable  conditions,  shows  us  this  respiration  to  be 
as  active,  and  even  more  so,  than  that  of  fishes.  This 
cannot  be  considered  as  only  a  curious  accessory  fact, 
of  which  we  must  take  but  slight  notice,  in  the  study 
of  the  biological  phenomena  of  yeast.  For,  d  priori,  it 
is  improbable  that  a  function  so  distinct,  so  sharply 
defined,  is  of  no  serious  importance.  On  the  other 
hand,  if  we  consider  what  is  passing  in  other  living 
beings,  from  the  highest  to  the  lowest  ranks  in  the  scale 


132 


ON  FERMENTATION. 


of  animal  and  vegetable  life,  we  see  respiration — that  is 
to  say,  combustion — at  the  expense  of  oxygen,  playing 
a  preponderating  part. 

Without  dwelling  on  the  animal  kingdom,  we  may 
remember  that  it  has  been  long  known  that  plants,  in 
darkness,  absorb  oxygen,  and  disengage  carbon  dioxide ; 
it  was  even  suspected,  with  good  reason,  that  this  re- 
spiratory function,  the  reverse  of  that  shown  by  parts 
exposed  to  the  sun,  was  independent  of  the  diurnal 
respiration,  and  that  it  belonged  to  another  class  of  cells 
containing  no  chlorophyll.  By  operating  on  immersed 
aquatic  plants,  we  can  demonstrate  this  fact  in  the 
clearest  and  most  elegant  manner.  Let  the  fresh  stalks 
of  Elodea  be  immersed  in  aerated  water,  of  which  the 
original  oxymetrical  degree  is  known.  The  flask,  com- 
pletely filled,  is  placed  in  the  dark.  At  the  end  of  two 
or  three  hours,  we  test  the  quantity  of  oxygen,  and 
we  find  a  diminution,  which,  as  was  the  case  with  the 
yeast,  at  equal  temperatures,  is  proportional  to  the 
quantity  of  the  plant,  and  the  duration  of  the  experi- 
ment, and  whose  absolute  amount  varies  with  the  tem- 
perature. 

If,  now,  we  warm  for  a  moment  the  water  and  the 
plant  to  about  50°  C.  (122''  R),  we  shall  destroy  for  ever 
the  activity  of  these  cells  containing  chlorophyll,  that  is 
to  say,  the  powers  which  the  plant  possesses  of  decom- 
posing carbon  dioxide  by  its  green  parts  ;  but  without 
lessening  its  activity  in  respiration  or  combustion.  We 
have  seen,  in  fact,  in  yeast,  that  this  function  is  only 
finally  modified  at  about  60°  C.  (140°  F.).  The  green 
parts  of  the  plant  are  dead,  but  it  is  still  capable  of  ful- 
filling certain  biological  functions.    The  flask  of  aerated 


FUNCTIONS  OF  YEAST. 


water  may  be  exposed  to  the  sun's  rays ;  when,  far 
from  observing  an  increase  in  the  quantity  of  dissolved 
oxygen,  the  opposite  is  seen. 

It  is  especially  in  the  parts  of  plants  which  have  to 
undergo  a  rapid  evolution,  and  a  marked  cellular  de- 
velopment, that  the  absorption  of  oxygen  and  internal 
combustion  show  themselves  to  be  very  active. 

Every  one  knows  that  in  the  germination  of  seed  or 
grain,  that  of  barley  in  the  manufacture  of  beer,  for 
instance,  the  internal  combustion  develops  a  considerable 
quantity  of  heat. 

The  blossoming  of  flowers  is  also  accompanied  by  a 
very  marked  oxidizing  respiration. 

Returning  to  yeast,  M.  Pasteur  has  shown  (Bullet, 
Soc.  Chimique,  p.  80,  1861)  that  beer-yeast,  sown  in  an 
albuminous  liquid,  such  as  the  water  of  yeast,  multiplies, 
even  when  there  is  no  trace  of  sugar  in  the  liquor,  pro- 
vided always  that  the  oxygen  of  the  air  be  present  in 
large  quantity  ;  deprived  of  air,  and  under  these  con- 
ditions, the  yeast  throws  out  no  buds.  The  same 
experiments  may  be  repeated  with  an  albuminous  liquid 
mixed  with  a  solution  of  unfermentable  sugar  such  as 
crystallized  sugar  of  milk ;  the  results  are  of  the  same 
^kind. 

The  yeast  formed  thus,  in  the  absence  of  sugar,  has 
not  changed  its  nature,  it  causes  sugar  to  ferment  if  it 
has  been  made  to  react  on  that  substance  without  the 
access  of  air.  We  must,  however,  remark,  that  the 
development  of  yeast  is  very  difficult  when  it  has  no 
fermentable  matter  to  nourish  it. 

On  the  other  hand,  this  same  observer  has  remarked 
that,  when  in  contact  with  air,  and  when  it  is  extended 


134 


ON  FERMENTATION. 


over  a  large  surface,  alcoholic  fermentation  is  more 
rapid  than  when  deprived  of  oxygen,  and  that  the 
budding  is  more  active,  since,  notwithstanding  the  great 
rapidity  of  the  fermentation,  the  relation  between  the 
newly-formed  yeast  and  the  decomposed  sugar  passes 
from  sV  to  i  or 

M.  Mayer  (Landw.  Versuchs.,  vol.  i6,  p.  290)  per- 
formed experiments,  from  which  it  would  appear  that 
oxygen  has  no  influence,  either  on  the  rapidity  of  the 
fermentation  or  on  the  quantity  of  newly-formed  yeast. 

However,  his  process  of  aeration  of  the  liquids  in 
fermentation,  which  consists  in  allowing  calcined  air  to 
pass  into  the  flask  three  times  a  day,  appears  to  me  to 
be  insufficient,  the  slowness  with  which  the  water  absorbs 
oxygen,  and  the  rapidity  with  which  yeast  absorbs  it, 
being  well  known ;  we  cannot,  then,  draw  from  the 
experiments  of  this  author  the  conclusions  unfavour- 
able to  Pasteur's  theory  which  he  drew  from  them. 

The  results  of  all  these  facts  are, — That  yeast,  like 
ordinary  plants,  buds  and  multiplies  even  in  the  absence 
of  fermentable  sugar,  when  it  is  furnished  with  free 
oxygen  ;  that  this  multiplication,  however,  is  favoured 
by  the  presence  of  sugar,  which  is  a  more  appropriate 
element  than  non-fermentable  hydrocarbon  compounds; 
and  also,  that  yeast  is  able  to  bud  and  multiply  in  the 
absence  of  free  oxygen,  but  that  in  this  case  a  ferment- 
able substance  is  indispensable. 

We  arc,  therefore,  compelled  to  arrive  at  the  con- 
clusion which  M.  Pasteur  has  drawn  from  all  these  facts; 
that  saccharine  matter  can  supply  free  oxygen  in  pro- 
portion to  the  yeast,  and  can  excite  it  to  bud.  M. 
Pasteur  has  gone  farther ;  he  thinks  that  the  ferment- 


FUNCTIONS  OF  YEAST. 


135 


ing  character  of  a  cell  is  due  to  the  power  which  it 
possesses  of  breathing  at  the  expense  of  sugar,  without 
the  contact  of  air,  and  that  the  decomposition  into 
alcohol  and  carbon  dioxide  is  the  consequence  of  a 
disturbance  of  equilibrium,  due  to  this  partial  abstrac- 
tion of  oxygen.  We  may  also  interpret  the  facts  in  the 
following  manner : — 

(1)  The  supply  of  oxygen,  and  the  combustions  to 
which  it  gives  rise  are  necessary  for  the  development 
and  the  reproduction  of  cell  life.  This  fact  is  abun- 
dantly established  for  all  the  forms  of  life  and  organs 
of  the  vegetable  kingdom. 

(2)  Yeast  possesses  the  power  of  resolving  the  sugar 
which  penetrates  by  endosmose  into  the  interior  of  the 
cell  into  alcohol,  carbon  dioxide,  glycerin,  succinic  acid, 
and  oxygen.  In  fact,  we  have  before  seen  (p.  23  of  the 
French)  that  M.  Monoyer  had  proposed  a  very  simple 
equation  to  represent  M.  Pasteur  s  formula  relative  to 
the  formation  of  succinic  acid  and  glycerin. 

In  this  equation  which  we  give  below, — 

4  (C12  Hn  On  +H  O),  or^ 
4(Ci2Hi2  0i2)  +  6H  O  =  Cg  H^Og  -h6  Q  Hg  Oe  +  2  C2  O4  +  Oo 

we  see  an  excess  of  oxygen  make  its  appearance  in 
the  second  member  of  the  equation,  and  M.  Monoyer 
adds,  "  we  may  suppose  that  this  oxygen  in  excess 
serves  for  the  respiration  of  the  yeast.  After  this,  the 
idea  is  not  without  foundation  that  fermentation  is  a 
primary  phenomenon,  due  to  a  special  action  of  the  yeast 
and  of  other  cells  ($echartier  and  Bellamy),  and  that, 

*  This  is  the  old  notation  form  of  given  by  M 
new  notation  form  will  be  found  on  the  page  referred  M 

f 


136 


ON  FERMENTATION. 


as  a  consequence  of  this  fermentation,  there  is  some 
oxygen  to  spare,  which  may  serve  the  purposes  of 
respiration,  and  consequently  may  promote  the  budding 
of  the  yeast. 

In  this  manner  of  explaining  the  facts,  the  yeast 
would  not  become  a  ferment  because  it  breathes  a  part 
of  the  oxygen  of  the  sugar  ;  but  it  may  breathe  a  part 
of  the  oxygen  of  the  sugar,  and  consequently  reproduce, 
precisely  because  it  sets  free  oxygen  by  decomposing 
the  sugar. 

Looking  at  the  question  in  a  more  general  point  of 
view,  we  may  also  say  that  respiratory  combustion  is 
to  the  living  organism  a  source  of  energy  necessary  for 
its  development.  In  the  decomposition  of  sugar  there 
would  be,  according  to  the  calculations  of  M.  Berthelot, 
a  disengagement  of  heat ;  the  quantity  of  heat  set  free 
in  this  phenomenon  would  be  about  i\  of  the  heat 
disengaged  by  the  complete  combustion  of  the  sugar 
decomposed.  (Comp.  Rend.,  vol.  59,  p.  901.)  In  this 
estimate  no  account  has  been  taken  of  the  heat  of 
solution  of  the  sugar  which  disappears,  nor  of  that  of 
the  solution  of  the  alcohol  formed,  positive  quantities 
which  would  tend  to  raise  the  heat  of  fermentation.  It 
is  not  even  necessary,  therefore,  to  have  recourse  to 
the  hypothesis  of  a  combustion  at  the  expense  of  the 
oxygen  of  the  sugar,  to  explain  how  the  phenomenon  of 
fermentation  can  feed  the  combustion,  and  become  a 
source  of  the  energy  indispensable  to  the  development 
of  the  plant. 

However  this  may  be,  there  is  an  evident  correlation, 
as  M.  Pasteur  observes,  bcwecn  fermentation  and  the 
development,  nutrition,  and  budding  of  yeast.    In  fcr- 


FUNCTIONS  OF  YEAST. 


mentation  without  oxygen,  the  relation  between  the 
new  yeast  and  the  decomposed  sugar  will  be  neces- 
sarily greater  than  in  fermentation  with  oxygen,  since 
in  the  former  case  the  budding  takes  place  only  under 
the  influence  of  the  oxygen  furnished  by  the  sugar,  and 
in  the  second,  of  that  furnished  both  by  the  medium 
and  the  sugar. 

We  ought  to  examine  what  takes  place  when  yeast  is 
left  to  itself,  without  nourishment  and  in  a  damp  state, 
without  the  intervention  either  of  saccharine  matter  or 
oxygen,  before  we  proceed  to  the  study  of  the  chemical 
modifications  produced  in  yeast,  while  it  is  placed  under 
conditions  favourable  to  its  nutrition  and  development. 
These  conditions,  as  we  have  seen,  return  always  to 
those  of  alcoholic  fermentation,  sugar  being  one  of  the 
indispensable  sources  of  nourishment  of  the  Saccharo- 
myces,  and  this  cannot  be  in  contact  with  it  without 
fermenting,  provided  that  the  other  conditions  of  nutri- 
tion are  fulfilled.  Does  it  preserve,  in  this  case,  its 
original  integrity,  without  any  change  in  the  chemical 
composition  of  its  immediate  principles }  In  other 
terms,  does  its  vitality  remain  in  a  latent  state,  to  mani- 
fest itself  afresh,  as  soon  as  sugar  or  oxygen  is  supplied.'^ 
A  prioriy  this  would  appear  improbable,  if  we  regard 
what  takes  place  in  the  tissues  of  plants.  In  fact, 
experiment  has  proved  that,  under  these  conditions,  the 
yeast  undergoes  important  modifications,  in  respect  of 
the  composition  of  its  organic  principles. 

This  question  has  been  studied  by  M.  Pasteur  first, 
and  then  successively  by  M.  Bechamp  and  by  the 
author  of  this  book. 

M.  Pasteur  having  set  5  grammes  of  sugar  to  fer- 
7  • 


138 


ON  FERMENTATION. 


ment  with  lo  grammes  ot  yeast,  in  the  form  of  paste, 
(2*155  grammes  of  dry  matter),  a  much  larger  quantity 
than  is  necessary  for  the  complete  decomposition  of  the 
sugar,  was  surprised  to  see  that  this  fermentation  did 
not  entirely  end,  that  it  had  a  tendency  to  continue  with 
a  weak  disengagement  of  gas,  and  when  Fehling's  liquor 
no  longer  revealed  in  it  the  slightest  trace  of  sugar. 
Having  arranged  in  receivers  reversed  over  mercury  the 
following  fermentations : — 

1st.  Sugar-candy  1*3 13 

Yeast  from  wine  (deposit  in  empty  casks)  6*950 
Pure  water  9*336 

2nd.  Sugar-candy  i'4425 

Yeast  from  beer  (2*1 5  grammes  when  dry)  lo'o 
Pure  water  9*210 

He  obtained,  in  two  days,  while  the  gaseous  disengage- 
ment was  still  sensible,  from — 

No.  I.  360  cubic  centimetres  (21*96  cub.  in.)  at  zero  (32°  F.),  and 

at  760  millimetres'  pressure  ; 
No.  2.  387 '5  cubic  centimetres  (23*6  cub.  in.)  at  the  same  tem- 


Of  carbon  dioxide,  entirely  absorbable  by  potass.  The 
theoretical  quantities,  even  when  a  deduction  is  made  of 
the  succinic  acid  and  glycerin — that  is  to  say,  those 
which  correspond  to  Gay-Lussac's  former  equation — 
those  which  give  the  highest  number  for  carbon  dioxide 
would  be — 


By  increasing  still  more  the  amount  of  yeast,  we 
arrive  at  more  conclusive  results. 


perature  and  pressure. 


No.  I. 
No.  2. 


341-8. 
375-5- 


FUNCTIONS  OF  YEAST. 


Thus,  '424  gramme  of  pure  sugar-candy,  made  to 
ferment  with  a  weight  of  damp  yeast  corresponding 
with  10  grammes  of  dry  matter,  furnished,  at  the  end 
of  two  days,  300  cubic  centimetres  of  carbon  dioxide  ; 
the  sugar  alone  could  only  furnish  no  cubic  centi- 
metres. 

The  liquid,  carefully  distilled,  gave  a  little  more  than 
0*6  gramme  of  absolute  alcohol.  The  weight  of  alcohol 
obtained  was  greater  than  the  whole  weight  of  the 
sugar  employed,  and  in  proportion  to  the  volume  of 
carbon  dioxide  formed. 

This  experiment  proves  that  when  yeast  is  mixed 
with  a  comparatively  very  small  weight  of  sugar,  after 
the  latter  has  been  decomposed,  the  activity  of  the 
yeast  continues,  reacting  on  its  own  tissues  and  its 
hydrocarbons  with  extraordinary  energy  and  rapidity, 
proceeding  more  and  more  slowly  as  the  process 
goes  on. 

If  we  put  an  end  to  the  fermentation  at  the  moment 
when  a  volume  of  carbon  dioxide  is  formed,  equal  or 
very  little  greater  than  that  which  corresponds  with  the 
weight  of  the  sugar  employed,  we  fiiid  no  more  sugar  in 
the  liquor.  This  observation  is  very  important,  because 
it  tends  to  prove  that  the  action  which  the  yeast  exerts 
on  its  own  elements  does  not  commence  till  it  is  de- 
prived of  sugar. 

It  is  not  necessary  to  fulfil  the  conditions  of  the 
preceding  experiments  (fermentation  with  excess  of 
yeast),  in  order  to  observe  fermentation  at  the  expense 
of  the  elements  of  the  yeast  itself;  it  is  sufficient,  as 
Pasteur  has  already  shown,  to  mix  fresh  beer-yeast  with 
water  at  25°  C.  (77 ''F.)  ;  we  soon  see  numerous  bubbles 


140 


ON  FERMENTATION. 


of  pure  carbon  dioxide  gas  rise,  and  it  is  easy,  by  dis- 
tillation, to  ascertain  the  production  of  alcohol.  Hydro- 
gen gas  and  signs  of  putrefaction  do  not  appear  till 
long  afterwards,  when  the  microscope  reveals  the 
presence  of  lactic  ferment  and  infusoria. 

M.  Bechamp,  who  took  up  this  question  after  M, 
Pasteur,  took  care  to  avoid  completely  the  formation 
of  infusoria,  by  employing  creosote  water ;  he  also 
ascertained  that  alcohol  and  carbon  dioxide  were 
produced  as  a  consequence  of  the  vital  activity  of  the 
yeast  when  without  nourishment  (deprived  of  oxygen 
and  sugar).  But  this  is  not  all,  for  this  curious  pheno- 
menon is  aUied  to  other  chemical  reactions  not  less 
interesting. 

It  is  well  known  that,  by  preserving  yeast,  in  the  form 
of  damp  paste,  in  a  hot  place  (25^^  to  30"  C,  77''  to 
86''  F.),  it  undergoes  considerable  softening,  and  entirely 
changes  its  appearance. 

This  modification  is  not  due  to  incipient  putrefaction. 
There  is  no  foundation  for  this  conclusion  ;  nothing  is 
formed  as  a  volatile  product,  except  carbon  dioxide  and 
alcohol.  The  microscope  reveals  no  organism  except 
the  SaccharomyceSy  especially  if,  as  M.  Bechamp  suggests, 
we  make  use  of  creosote  water.  If,  now,  we  treat  this 
softened  yeast  with  luke-warm  water,  we  can  extract 
from  it  a  much  greater  quantity  of  soluble  and  diffu- 
sible principles  than  by  employing  fresh  yeast. 

Thus,  100  grammes  of  fresh  yeast,  leaving  after  des- 
sication  at  100*^  C.  (212°  F.),  a  fixed  residue  of  30 
grammes,  give  after  washing,  before  the  softening  takes 
place,  only  23  grammes  of  dry  residue.  The  loss  by 
washing  is  8  per  cent. 


FUNCTIONS  OF  YEAST. 


The  same  yeast,  softened  spontaneously  for  two  days, 
and  then  washed,  leaves  a  residue  weighing,  when  dry, 
14  grammes  ;  the  loss  by  washing  is  thus  16  grammes. 

Yeast  has  therefore,  by  softening,  transformed  into 
soluble  principles  8  grammes  of  previously  insoluble 
principles,  per  cent,  of  damp  yeast,  or  26*66  grammes 
per  cent,  of  dry  yeast.  During  this  internal  reaction 
of  yeast,  when  kept  damp,  and  without  nourishment, 
M.  Bechamp  observed  the  production  of  pure  nitrogen. 

The  water  by  which  it  has  been  washed  contains 
acetic  acid,  a  perceptible  portion  of  soluble  alterative  fer- 
ment (zymase,  of  which  we  shall  presently  speak  ;  ^^^the 
chapter  on  soluble  ferments),  an  albuminoid  principle, 
soluble,  coagulableby  heat,  and  nearly  allied  to  albumin, 
from  which  it  differs  by  rotating  power  ;  a  gummy  matter, 
which  nitric  acid  transforms  into  mucic  acid,  very  like 
arabin,  from  which,  however,  it  differs  by  rotating  power  ; 
we  find,  also,  tyrosine  and  leucine,  and  an  uncrystallizable 
sirupy  matter,  alkaline  and  alkaline-earthy  phosphates 
in  observable  proportions. 

Resuming  the  question,  the  author  confirmed  the 
results  obtained  by  M.  Bechamp,  and  added  to  them 
new  facts.  The  extract  of  softened  yeast  contains, 
besides  the  principles  mentioned  above,  nitrogenous 
compounds  of  the  sarcine  group,  which  had  not  hitherto 
been  noticed  in  the  vegetable  economy. 

The  following  is  the  analytical  process  which  I  have 
followed.  The  yeast,  digested  without  the  addition  of 
any  nutritive  principles,  is  boiled  with  a  considerable 
quantity  of  water  to  coagulate  the  albumin  ;  it  is  then 
filtered.  The  liquid,  which  has  a  slightly  acid  reaction, 
is  concentrated  in  a  water  bath  to  a  sirupy  consistence  ; 


142 


ON  FERMENTATION. 


it  assumes,  as  it  grows  cold,  a  gelatinous  crystallized 
form  composed  of  small  crystals  forming  a  kind  of 
paste  in  a  brownish  syrup.  The  whole,  placed  in  a 
flask,  is  boiled  for  some  time  with  a  great  excess  of 
strong  alcohol  (92  per  cent.)  ;  a  deep  coloured  pitchy 
mass  is  separated,  which  sticks  to  the  sides  of  the 
flask.  The  alcoholic  solution,  when  properly  concen- 
trated, furnishes,  as  it  grows  cool,  an  abundant  crystal- 
line deposit,  which,  after  filtration,  washing  with  cold 
alcohol,  and  pressure,  is  almost  w^hite.  This  crystallized 
mass,  formed  of  very  thin  flakes  or  hyaline  globules,  as 
well  as  that  which  is  obtained  by  concentrating  the 
alcoholic  mother-liquor,  is  almost  exclusively  formed  of 
sulphuretted  psetido-leitcine,  with  a  little  tyrosine.  The 
mother-liquor  separated  at  the  second  crystallization 
is  distilled  in  a  water-bath  to  drive  off*  the  alcohol. 
The  remainder  is  diluted  with  water,  and  solution  of 
barytes  is  added  to  precipitate  the  phosphates.  The 
excess  of  barytes  is  removed  from  the  filtered  liquid  by 
a  current  of  carbon  dioxide.  The  filtered  liquid  is  then 
boiled  with  an  excess  of  cupric  acetate.  A  brownish 
flaky  precipitate  is  formed,  which  contains  carnine, 
sarcine,  xanthine,  and  guanine,  combined  with  copper 
oxide.  The  filtered  liquor  above  this  precipitate,  which 
is  comparatively  not  very  abundant,  yields,  when  the 
copper  has  been  removed  by  sulphuretted  hydrogen, 
and  it  has  been  concentrated,  a  crystalline  mass,  from 
which  cold  alcohol  extracts  a  sirupy  nitrogenous  sub- 
stance, of  a  sweet  taste,  leaving  a  crystallized  mixture 
of  leucine  and  butalinine. 

The  cupric  precipitate,  produced  by  boiling  in  cupric 
acetate,  is   thoroughly  washed  with  hot  water,  then 


FUNCTIONS  OF  YEAST. 


treated,  by  the  warm  process,  with  dilute  hydrochloric 
acid.  Nearly  all  of  it  is  dissolved,  with  the  exception 
of  black  flakes  of  copper  sulphuret.  The  solution, 
filtered  while  warm,  deposits,  when  cold,  a  large  part 
of  the  copper  combination,  which  had  been  dissolved. 

This  deposit,  washed  and  decomposed  by  sulphuretted 
hydrogen,  furnishes  carnine,  which  is  purified  by  being 
crystallized  in  water,  being  deprived  of  colour,  if  neces- 
sary, by  a  little  animal  charcoal.  The  hydrochloric 
mother-liquor  which  deposited  the  copper  combination 
of  carnine,  having  been  deprived  of  its  copper  by 
sulphuretted  hydrogen,  and  then  concentrated,  gives 
first  crystals  of  xanthine  hydrochlorate  ;  then,  by  con- 
centrating the  decanted  liquor  in  the  cold,  crystals  ot 
guanine  hydrochlorate,  from  which  the  guanine  is  ex- 
tracted by  precipitating  it  with  an  excess  of  ammonia, 
which  dissolves  the  xanthine,  and  leaves  the  guanine. 

The  sarcine  is  obtained  by  precipitating  by  ammonia 
and  silver  nitrate  the  nitric  solution  of  the  first  copper 
precipitate.;  by  washing  with  water  of  ammonia  the 
dirty-white  gelatinous  precipitate  which  is  formed,  and 
then  crystallizing  it  in  boiling  nitric  acid  at  12°  Baume  ; 
we  thus  obtain  immediately  the  nitro-argentine  combi- 
nation of  sarcine.  This  is  decomposed  by  sulphuretted 
hydrogen  ;  the  liquid,  when  filtered,  and  concentrated, 
has  ammonia  added  to  it,  which,  by  concentration, 
leaves  the  sarcine  in  fineneedle-like  crystals. 

The  pitchy  precipitate  formed  at  first,  by  the  addition 
of  alcohol  to  the  concentrated  extract  of  digested  yeast, 
is  in  great  part  formed  of  earthy  phosphates,  tyrosine, 
and  gummy  matter. 

The  leucine  extracted  from  the  yeast  after  its  diges- 


144 


ON  FERMENTATION. 


tion  shows  a  peculiarity  noticed  by  Hesse.  It  contains 
a  quantity  of  sulphur,  which  varies  within  certain  Hmits, 
and  may  attain  to  4  per  cent.  My  analyses,  made  from 
very  pure  products,  crystallized  several  times  in  alcohol, 
and  having  the  appearance  of  beautiful  white  nacreous 
flakes,  gave  from  2*93  to  2*14  per  cent,  of  sulphur.  This 
cannot  be  eliminated  by  long  boiling  at  100°  C.  (212°  F.) 
with  a  mixture  of  potass  and  lead  hydrate.  The 
hydrogen  of  these  leucines  have  always  been  found 
to  be  somewhat  deficient  (9*34  or  9*6  per  cent,  instead 
of  9-9). 

It  appears  that  the  sulphur  forms  an  integral  part  of 
the  molecule,  and  is  not  found  in  the  state  of  a  sulphu- 
retted body  mixed  with  leucine.  At  any  rate,  repeated 
and  very  careful  purifications  do  not  succeed  in  splitting 
up  this  mixture. 

Let  us  now  see  what  may  be  the  signification  of  these 
results. 

All  the  nitrogenous  compounds  noticed  as  being  the 
products  of  spontaneous  changes  in  the  yeast  have 
been  obtained  directly  by  the  splitting  up  of  albumin 
and  albuminoid  substances.  {See  the  chapter  on  these 
bodies.)  Their  origin  is,  therefore,  not  doubtful  ;  they 
are  formed  by  the  decomposition  of  certain  -  insoluble 
proteids  in  the  yeast,  and  by  a  chemical  process  similar 
to  that  which  takes  place  in  the  animal  tissues  ;  for  it  is 
impossible  to  mistake  the  great  analogy  of  composition 
which  exists  between  the  extract  of  digested  yeast  and 
extracts  obtained  from  animal  tissues.  This  chemical 
phenomenon  is  also  comparable  to  those  which  are 
observed  by  causing  dilute  warm  sulphuric  acid,  or  the 
alkalis,  such  as  potash  and  barytes,  to  react,  under  ccr- 


FUNCTIONS  OF  YEAST. 


tain  conditions,  on  proteids  ;  we  thus  obtain,  in  fact, 
simifar  products. 

Albumin,  obtained  in  such  large  proportions  in  the 
extract  of  digested  yeast,  must  be  one  of  the  results  of 
the  splitting  up  ;  it  is  probable  that  yeast  has  no  action 
on  the  one  substance  among  proteids  which  resists  so 
strongly  chemical  agents. 

This  opinion  is  corroborated  by  the  interesting  obser- 
vation of  M.  Gautier,  who  proved  that  fibrin  splits  up, 
under  the  influence  of  salt  water,  into  albumin  and 
another  albuminoid  principle.  Zymase,  or  alterative 
ferment,  which  exists  in  large  proportions  in  the  extract 
of  softened  yeast,  also  represents  one  of  the  substances 
derived  from  the  decomposition  of  insoluble  proteids. 

I  have  before  said  that  we  succeed,  by  direct  analysis, 
in  separating  an  uncrystallizable  nitrogenous  principle 
of  a  sweet  taste.  This  substance  resembles  in  its  pro- 
perties the  hemiproteidin  on  hemialbumin  formed  by 
the  action  of  boiling  dilute  sulphuric  acid  on  albumin. 

As  to  the  tyrosine,  leucine,  butalanine,  the  sarcinic 
bases,  is  evident  that  they  are  the  directly  derived  pro- 
ducts of  albuminoid  substances. 

With  respect  to  the  proceeds  of  the  sugar  which 
furnish  the  alcohol  and  carbon  dioxide  of  the  spontaneous 
fermentation  of  yeast,  and  of  the  gummy  matter,  their 
source  has  not  been  ascertained  with  as  great  certainty. 

It  is  generally  admitted,  in  accordance  with  the  views 
of  Payen  and  Schlossberger,  that  washed  yeast  is  com- 
posed of  cellulose  and  insoluble  proteids. 

The  question  whether  the  sugar  and  gum  which  show 
themselves  in  notable  proportions  in  the  extract  of  yeast, 
when  digested  and  washed,  ought  to  be  considered  as 
the  products  of  the  physiological  decomposition  of  the 


ON  FERMENTATION. 


albuminoid  substances,  or  the  results  of  a  transforma- 
tion of  cellulose,  can  be  decided  only  by  a  series  of  quan- 
titative experiments.  The  following  results  give  only 
an  approximate  solution,  but  they  prove,  at  least,  that 
the  greater  part,  if  not  the  whole,  of  the  solid  principles 
contained  in  the  extract  of  yeast  when  washed  in  cold 
water  and  digested,  must  be  derived  from  proteids. 

ICO  grammes  of  fresh  yeast  contain  30  grammes  of 
solid  matter.  This  includes  9*28  per  cent,  of  nitrogen  ; 
I  whence  it  results,  that  100  grammes  of  fresh  yeast 
1  contain  278  grammes  of  nitrogen. 

ICQ  grammes  of  fresh  yeast,  w^ashed  with  boiling 
water  until  it  runs  off  clear,  leave  20  grammes  of  solid 
matter.  This  contains  10*17  per  cent,  of  nitrogen. 
The  result  is  that  100  grammes  of  yeast,  washed  with 
boiling  water,  contain  2*03  grammes  of  nitrogen. 

100  grammes  of  yeast,  digested  for  fifteen  hours  at 
35°  C.  (95°  F.),  and  then  washed  with  boiling  water, 
contain  12*5  grammes  of  solid  matter,  yielding  7*55  per 
cent  of  nitrogen. 

100  grammes  of  yeast,  digested  and  then  washed 
with  boiling  water,  contain,  then,  70*94  grammes  of 
nitrogen.  The  loss  of  nitrogen  arising  from  the  diges- 
tion and  washing  is  r82  —  0*75  =:  1*07. 

The  loss  of  solid  products  due  to  digestion  and  sub- 
sequent washing  amounts  to  17*5  —  10  :=:  7*5. 

But  proteids  contain,  on  an  average,  15 '5  per  cent,  of 
nitrogen  ;  ro7  gramme  of  nitrogen  correspond  to  6*9 
grammes  of  proteinic  matter.  Consequently,  out  of  7*5 
grammes  of  the  principles  which  have  become  soluble  by 
the  process  of  digestion,  6*g  grammes  or  are  derived 
from  albuminoid  substances. 

On  the  other  hand,  a  direct  quantitative  estimate  of 


FUNCTIONS  OF  YEAST. 


the  nitrogen  in  the  dry  extracts  gave  1 2*5  per  cent,  of 
nitrogen.  It  is,  then,  evident  that  the  yeast  must  have 
given  up  part  of  its  non-nitrogenous  elements  ;  this  last 
calculation  would  lead  us  to  the  proportion  of  ^  proteids, 
and  ^  hydrocarbon  matter,  if  we  take  no  account  of 
the  elements  of  water  which  are  necessarily  united  with 
the  albuminoid  matter  at  the  time  of  its  splitting  up 
and  transformation  into  bodies  such  as  leucine.  By 
making  an  approximate  estimate  of  this  water  of  hydra- 
tation,  we  should  find  -f-f  of  albuminoid  and  of 
hydrocarbon  matter.  It  is  not  very  probable  that  the 
gum  has  an  albuminoid  origin;  it  could  at  most  be 
derived  only  from  the  decomposition  of  a  substance 
analogous  to  tunicin  or  chitin. 

The  principles  yielded  to  water  by  fresh  yeast  in  a 
much  smaller  proportion  are  of  the  same  nature  as 
those  which  we  have  found  before  ;  it  cannot,  indeed,  be 
otherwise.  The  conditions  under  which  M.  Bechamp 
and  I  carried  on  our  researches  necessarily  exaggerated 
the  effects  of  a  continuous  cause.  As  soon  as  the  yeast 
finds  no  more  sugar,  it  reacts  on  its  own  elements,  and  it 
is  difficult  to  find  yeast  which  has  not  been  placed  in  this 
situation  for  a  longer  or  shorter  time.  It  is  probable 
that  all  kinds  of  yeast  give  reactions  of  this  nature  in 
the  fermenting  vats,  when  the  sugar  begins  to  fail.  M. 
Bechamp  says  that  he  has  found  tyrosine  and  leucine  in 
the  aqueous  extract  of  all  fermentations  which  he  has 
examined  ad  hoc.  Before,  therefore,  we  draw  too  positive 
conclusions,  we  must  know  whether  the  fermentations 
have  been  studied  immediately  after  the  total  decompo- 
sition of  the  sugar. 

M.  Pasteur  has  shown  us  that  spontaneous  fermenta- 
tion does  not  appear  till  this  moment ;  it  would  be 


148 


ON  FERMENTATION. 


interesting  to  ascertain  whether  the  formation  of  nitro- 
genous excremental  substances,  such  as  leucine  or  tyro- 
sine, takes  place  during  fermentation  or  no  ;  we  should 
thus  see  whether  this  phenomenon  is  allied  to  spon- 
taneous fermentation,  or  is  independent  of  it. 

M.  Bechamp,  considering  the  spontaneous  fermenta- 
tion of  yeast  and  its  concomitant  phenomena  (the 
production  of  acetic  acid,  tyrosine,  &c.)  has  given  a 
physiological  theory  of  fermentation  which  may  be 
shortly  stated  thus : — 

Yeast,  like  every  living  organism,  shows  phenomena 
of  two  kinds  ;  those  of  nutrition  and  assimilation,  which 
are  subordinate  to  the  presence  of  its  nutritious  princi- 
ples (sugar,  nitrogenous  compounds,  mineral  salts). 
These  various  principles,  penetrating  by  endosmose 
into  the  cell,  undergo  there  suitable  transformations, 
and  are  converted  into  tissues  of  recent  formation  in 
the  new  cells  which  are  formed  by  budding.  Together 
with  these  phenomena  of  nutrition,  and  side  by  side 
with  them,  other  inverse  reactions,  those  of  disassimila- 
tion,  take  place,  by  which  the  tissues  are  changed  into 
excrementitious  products,  unsuited  to  the  life  of  the  cell, 
and  these  are  eliminated.  The  production  of  carbon 
dioxide  and  of  alcohol  are  the  consequences  of  this 
process,  and  belong  to  disassimilating  reactions.  In  this 
theory,  M.  Bechamp  develops  the  ideas  of  M.  Dumas 
as  to  the  part  played  by  yeast  and  other  ferments.  It 
is  certain  that  the  production  of  carbon  dioxide  and 
alcohol  without  sugar,  at  the  expense  of  the  constituent 
elements  of  the  yeast  itself,  gives  some  support  to  the 
opinion  of  15echamp,  whether  or  no  we  admit  that  this 
production  is  the  result  of  the  formation  of  buds,  in  which 
the  new  cells  are  nourished  at  the  expense  of  the  old  ones. 


FUNCTIONS  OF  YEAST. 


149 


On  the  whole,  this  theory  throws  but  Httle  light  on 
the  question  relating  to  the  very  essence  of  the  pheno- 
menon. Whether  the  sugar  is  decomposed  with  or  with- 
out previous  assimilation  is  of  little  importance  ;  we  are 
no  nearer  knowing  why  it  is  decomposed,  and  scarcely 
any  one  now  doubts  that  the  decomposition  of  sugar 
is  a  biological  phenomenon.  We  must  now  examine 
some  special  points  relating  to  alcoholic  fermentation. 

It  has  been  long  thought  that  alcoholic  fermentation 
could  take  place  under  two  distinct  circumstances,  ac- 
cording as  yeast  is  added  to  a  solution  of  pure  sugar  in 
water,  or  to  sugared  water  containing  the  nitrogenous 
and  saline  principles  necessary  for  its  nourishment.  In 
the  first  case,  it  was  thought  that  the  ferment  acts  with- 
out reproducing  itself ;  while  in  the  second  case,  which  is 
that  occurring  in  the  brewing  of  beer,  it  acts  and  repro- 
duces itself.  More  than  this,  Thenard  had  observed 
that,  in  the  fermentation  of  20  parts  of  yeast  and  ICQ 
parts  of  sugar,  there  only  remain,  after  all  the  dis- 
engagement of  carbon  dioxide  has  ceased,  137  parts  of 
an  insoluble  residue,  which  may  be  reduced  even  to  10 
by  fresh  contact  with  sugar.  Thus  yeast,  while  exciting 
the  fermentation  of  pure  sugar,  partially  destroys  itself 

M.  Pasteur,  on  the  contrary,  admitting  it  to  be 
proved  by  his  experiments  that  the  budding  and  multi- 
plication of  yeast  are  phenomena  which,  in  a  constant 
manner,  accompany  all  alcoholic  fermentation ;  explains 
the  formation  of  new  globules  in  a  solution  of  pure 
sugar  in  water  by  nutrition  at  the  expense  of  the  nitro- 
genous soluble  substances  of  the  original  yeast.  In 
this  case,  whatever  soluble  nitrogenous  aliment  there  is 
in  the  ferment  employed  becomes  fixed  in  an  insoluble 
state  in  the  newly-formed  globules.    It  is  certain  that 


ON  FERMENTATION. 


yeast  formed  in  a  suitable  medium,  such  as  the  wort  of 
beer,  is  gorged  with  soluble  nitrogenous  principles,  which 
it  can  yield  to  water,  and  which  are  eminently  suited  to 
the  nutrition  of  new  yeast. 

In  order  to  remove  all  doubt  as  to  the  reality  of  this 
explanation,  it  w^as  necessary  for  Pasteur  thoroughly  to 
explain  away  those  of  Thenard's  results,  quoted  above, 
which  seem  to  contradict  his  opinion. 

The  following  table  sums  up  his  observations  on  the 
fermentation  of  pure  sugar  with  yeast  : — 


Weight  of  pure  sugar- 
candy. 

Weight  of  yeast,  washed  while 
fresh,  in  the  state  of  a  soft 
paste. 

Weight  of  the  yeast  when 
1       dried  at  100°  C.  (212°  F.). 

Weight  of  yeast  deposited 
.   after     fermentation,  when 
dried  at  100°  C.  (212°  F,). 

Weight  of  extract,  the  soluble 
part  of  the  yeast  remaining 
(     in  the  fermented  liquid,  and 
insoluble  in  the  mixture  of 
alcohol  and  ether. 

Sum  of  the  weights  of  the 
yeast   deposited  after  fer- 
mentation and  of  the  so- 
luble extractive  part  remain- 
ing in  the  fermented  liquor. 

Excess  of  this  sum  over  the 
weight  of  yeast  exposed  to 
fermentation. 

gl-. 

gr- 

gr- 

gr. 

gr. 

A 

TOO 

20  000 

4-626 

3  "230 

2-320 

5-550 

0934 

B 

50 

lO'OOO 

2-213 

2001 

0819 

2820 

0-407 

C 

100 

i6*ooo 

4-604 

4-385 

not  deter- 
mined. 

D 

100 

I0"000 

2-313 

2-486 

I -080 

3-566 

1-253 

E 

100 

13700 

2-626 

2-965 

0-964 

3-929 

I -303 

F 

100 

6-254 

1-198 

1-700 

0-631 

1-331 

1-133 

G 

16 

3'i59 

0-699 

0-712 

not  deter- 
mined. 

II 

4 

'i  '474 

0-326 

0335 

I 

20 

1-878 

0-476 

0590 

0133 

0-723 

0'24.7 

FUNCTIONS  OF  YEAST. 


We  see  that  in  the  experiments  A  B  C,  in  which  the 
weights  of  yeast  in  the  form  of  paste  is  above  15  or  20 
per  cent,  of  the  weight  of  sugar,  after  fermentation  less 
yeast  is  collected  than  is  originally  put  in  ;  these  are, 
therefore,  precisely  the  conditions  of  Thenard's  experi- 
ments, who  employed  and  recommended  20  parts  of 
yeast  in  form  of  paste  to  100  of  sugar. 

If,  on  the  contrary,  the  weight  of  yeast  in  the  form 
of  paste  is  equal  to  or  less  than  10  per  cent,  of  the 
sugar,  more  yeast  is  collected  than  had  been  first 
employed  (exper.  D  E  F  G  H  I). 

And,  in  every  case,  by  adding  the  weight  of  extractive 
matter  to  that  of  the  yeast  remaining  after  fermentation 
(having  deducted  the  glycerin  and  succinic  acid  which 
are  in  solution  in  the  fermented  liquor,  and  come  from 
the  yeast),  we  find  that  the  sum  always  exceeds  very 
sensibly  the  whole  weight  of  the  original  yeast. 

The  increase  amounts  to  about  i*2  or  i'5  per  cent,  of 
the  weight  of  the  sugar  decomposed. 

The  result  of  this  appears  to  be,  that  in  Thenard's 
experiments,  the  disappearance  of  yeast  was  only 
apparent.  Less  was  collected  because  much  was  used, 
and  because  the  soluble  part  which  passes  into  the  liquid 
is  greater  than  the  weight  of  the  newly-formed  globules. 

M.  Duclaux  (Theses  de  la  Faculte  des  Sciences  de  Paris, 
1865)  arrived  at  results  of  the  same  kind. 

In  ICQ  grammes  of  fermented  sugar  he  found — 


Dry  yeast  employed. 


Dry  yeast  obtained. 
14*02 


17-32 

8-66 


8-51 
478 
2-53 


4*33 

2*l6 


152 


ON  FERMENTATION. 


However,  in  other  trials  made  by  the  same  author, 
the  weight  of  yeast  found  exceeded  that  of  the  yeast 
employed,  even  in  the  case  in  which  the  original  pro- 
portion exceeded  i6  per  cent,  of  sugar.  This  difference 
is  attributed  by  the  author  to  the  use  at  first  of  a  kind 
of  yeast  poorer  in  extractive  principles,  and  yielding 
fewer  solubler  substances  to  the  water. 

The  results  obtained  by  Pasteur  and  Duclaux,  in 
fermentation  conducted  on  a  small  scale,  have  been 
confirmed  by  other  experimentalists,  such  as  those  of 
Mayer,  Fitz. 

From  the  whole  results  published,  it  seems  that  we 
may  conclude  that,  within  certain  limits,  and  with  some 
variations,  the  weight  of  the  newly-formed  yeast  is  in 
proportion  to  the  weight  of  the  sugar  decomposed,  and 
equal  to  from  i*5  to  3  per  cent,  of  the  fermented  sugar. 

We  ought,  however,  to  add,  that  the  information 
afforded  by  commercial  experience,  by  the  manufac- 
turers of  compressed  yeast,  do  not  agree  with  these 
conclusions. 

According  to  Marcher  and  Schulze,  we  are  able  to 
obtain  28*6  parts  of  dry  yeast  for  100  of  alcohol,  or, 
which  comes  to  the  same  thing,  14*66  parts  of  dried 
yeast  for  100  parts  of  decomposed  sugar. 

In  the  fermentation  of  the  must  of  grapes,  the  forma- 
tion of  yeast  seems  much  greater  in  proportion  to  the 
fermented  sugar  than  M.  Pasteur's  ratio  would  make  it. 

M.  Pasteur,  who  carries  on  with  so  much  success  the 
manufacture  of  pure  yeast  on  a  large  scale,  by  the  fer- 
mentation of  the  wort  of  beer  in  a  close  vessel,  obtains 
1*5  kilogrammes  (3*3  lbs.  avoir.)  of  expressed  yeast,  which 
contains  30  per  cent,  of  dry  matter,  say  450  grammes 


FUNCTIONS  OF  YEAST. 


(•99  lbs.  avoir.)  of  dried  yeast  per  hectolitre  (22  gallons) 
of  beer  ;  allowing  that  this  beer  contains  8  per  cent,  of 
alcohol,  fermentation  would  have  acted  on  15*5  kilo- 
grammes of  sugar,  there  would  then  be  2*9  grammes  of 
new  yeast  for  100  of  decomposed  sugar ;  these  numbers 
do  not  differ  so  much  as  the  first,  from  the  results 
obtained  on  a  small  scale  ;  it  is  true,  however,  that  in  this 
calculation,  we  have  taken  for  the  proportion  of  alcohol 
a  very  high  weight  which  ought,  perhaps,  to  be  reduced. 

The  apparent  contradiction  between  the  results  of  the 
laboratory  experiments  and  those  of  commercial  opera- 
tions, may  be  explained  by  admitting,  that  the  develop- 
ment and  multiplication  of  the  cells  of  yeast  are  not  so 
much  dependent  on  the  quantity  of  sugar  decomposed, 
as  the  laboratory  experiments  seem  to  indicate,  that 
they  may  vary  within  very  wide  limits,  according  to  the 
composition  of  ihe  medium  in  which  the  operation  takes 
place. 

This  explanation  seems  to  be  corroborated  by  the 
following  experiments  of  Mayer  (Landwi,  Versuchsz, 
vol.  16,  p.  304). 

The  author  fermented  two  portions  of  the  boiled 
must  of  grapes  of  the  capacity  of  190  cubic  centimetres 
(i  1*59  cub.  in).  To  one,  he  added  imperceptible  traces  of 
wine-lees  {Saccharomyces  ellips\  and  sowed  in  the  other 
a  little  beer-yeast.  When  the  sugar  had  disappeared, 
which  required  several  weeks,  he  found — 

ALCOHOL  CORRESPONDING  YEAST 
FORMED.  SUGAR  DECOMPOSED.  FORMED. 

gr.  gr.  gr. 

1.  Saccharomyces  ellip.     .    13-11  25*6  2*38 

2.  cerevisicE    152  297  2*04 


154 


ON  FERMENTATION. 


Therefore  for  lOO  parts  of  sugar  decomposed  there  were 
formed — 

No.  I.  9*2  of  yeast. 
No.  2.  6-8  „ 

numbers  much  higher  than  the  results  obtained  in 
fermentations  under  other  conditions  of  the  medium. 

On  the  other  hand,  M.  Pasteur  found  that  in  fer- 
mentations, in  the  presence  of  nitrogenous  and  mineral 
nutritive  matter,  nearly  i  per  cent,  of  the  weight  of 
sugar  was  formed  in  yeast  and  soluble  products ;  a  little 
less,  therefore,  than  when  we  operate  with  yeast  com- 
pletely formed,  and  pure  sweetened  water. 

Thus,  to  9*899  grammes  of  pure  sugar-candy,  dissolved 
in  a  sufficient  quantity  of  water,  were  added  20  cubic 
centimetres  of  boiled  and  filtered  water  with  which  fresh 
yeast  had  been  washed  (containing  0*334  grammes  of 
albuminoid  and  mineral  matter),  it  was  then  filled  up  to 
100  cubic  centimetres  and  a  trace  of  fresh  yeast  added. 
The  sugar  had  disappeared  at  the  end  of  eleven  days ; 
0*152  grrammes  of  dry  yeast  were  collected.  The  nitro- 
genous remainder  left  after  the  evaporation  of  the  liquid, 
when  the  glycerin  and  succinic  acid  had  been  removed, 
weighed  0*260  grammes. 

Thus,  o*334  grammes  of  nutritive  matter  were 
employed,  and  0*1 52  grammes  of  yeast  +  0*260  grammes 
of  soluble  nitromineral  matter  was  found  ;  in  all,  0*412 
grammes.    The  difference  is  0*78  grammes. 

When  pure  sugar  ferments  with  a  limited  quantity  of 
yeast,  this  becomes  exhausted,  and  at  last  becomes  unfit 
to  effect  fresh  decompositions  of  sugar,  in  the  absence 
of  soluble  nutritive  materials.  This  ought,  in  fact,  to 
be  the  case.     The  amount  of  the  nutritive  materials 


FUNCTIONS  OF  YEAST. 


which  are  soluble,  or  become  so  during  the  fermentation, 
and  which  are  contained  in  the  limited  yeast,  being 
gradually  utilized  for  the  formation  of  fresh  globules, 
there  ought  to  come  a  time  when  the  yeast  will  be 
destroyed,  for  want  of  nutriment.  This  phenomenon 
of  the  nutrition  of  the  yeast  at  its  own  expense,  does 
*  not  exclude  the  presence  of  nitrogenous  matter  in  the 
no  longer  fermenting  saccharine  liquid,  for  it  may 
happen  that  a  part  of  the  nitrogenous  principles  elimi- 
nated by  the  yeast  are  unsuited  to  its  nourishment,  and 
only  represent  excremental  products.  Leucine  and 
tyrosine  seem  to  belong  to  this  order  of  products ;  in 
fact,  some  direct  experiments  of  Mayer  have  proved 
that  they  are  unsuited  to  development. 

In  a  fermentation,  in  which  1*198  grammes  of  washed 
yeast  (the  weight  of  dried  matter,  containing  977 
grammes  per  cent,  of  nitrogen)  had  been  employed  to  fer- 
ment ICQ  grammes  of  sugar,  1745  grammes  of  yeast  were 
collected  after  fermentation ;  this  contained  5 '5  percent,  of 
nitrogen  ;  the  extractive  residuum  of  the  fermented  liquid, 
when  deduction  had  been  made  of  the  succinic  acid  and 
glycerin,  weighed  o*6  grammes  and  contained  3*8  per 
cent,  of  nitrogen.  Under  these  conditions,  the  yeast 
was  exhausted — that  is  to  say,  the  soluble  nitrogenous 
principles  contained  in  the  liquid  had  become  unfit  for  the 
development  of  new  cells,  and  represented  excremental 
products.  We  know  that  among  the  products  left  undeter- 
mined by  M.  Pasteur,  we  must  place  leucine  and  tyrosine. 

This  experiment  shows  us  the  cause  of  the  diminu- 
tion of  nitrogen  in  the  yeast  after  fermentation.  On  the 
one  hand,  the  total  weight  of  yeast  has  increased  by  the 
addition  of  principles  (cellulose)  furnished  by  the  sugar, 


ON  FERMENTATION. 


and  not  nitrogenous  ;  on  the  other  hand,  the  original 
yeast  has  yielded  to  the  liquid  soluble  nitrogenous  pro- 
ducts which  have  become  unfit  for  its  nutrition.  Thus 
— the  apparent  disappearance  or  diminution  of  the  nitro- 
gen of  the  yeast  during  fermentation  with  pure  sugar 
is  explained  in  a  very  natural  manner,  a  phenomenon 
which  had  so  puzzled  Thenard,  and  which  Dobereiner  * 
had  wrongly  thought  might  be  attributed  to  the  for- 
mation of  ammoniacal  salts.  We  have  already  seen  by 
Pasteur's  very  careful  experiments,  that  the  ammonia 
of  the  liquid  has  a  tendency  rather  to  disappear  than  to 
increase  during  the  fermentation. 

According  to  Dubrunfaut  (Comp.  Rend.,  July,  1 871), 
the  wort  of  beer,  which,  under  ordinary  circumstances, 
reproduces  sevenfold  the  weight  of  the  yeast  employed, 
is  so  rich  in  matter  reproductive  of  ferment,  that  it  is  far 
from  being  exhausted  by  this  process.  An  addition  of 
sugar  excites  an  increase  proportional  to  the  sugar 
supplied.  If  it  has  not  been  employed  in  excess,  the 
normal  condition  of  the  ferment  with  respect  to  nitrogen 
has  not  changed  ;  if  the  contrary  has  been  the  case,  the 
amount  of  nitrogen  lies  between  that  of  fertile  and  of 
barren  yeast.  All  kinds  of  yeast,  the  results  of  fermen- 
tation other  than  that  of  beer  made  from  malt,  are  in 
this  condition  ;  they  give  an  amount  of  nitrogen  varying 
between  O'lo  and  '05. 

Beer-yeast,  notwithstanding  the  loss  of  weight  sus- 
tained in  washing,  preserves  its  proportion  of  nitrogen, 
while  its  richness  in  salts  decreases  from  'lo  to  '02, 
However  often  it  may  be  washed,  we  can  never  obtain 
water  quite  free  from  albuminoid  and  saline  matter;  which 
proves  that  the  yeast  continues  to  live,  even  in  pure  water, 


FUNCTIONS  OF  YEAST. 


157 


and  to  exercise  there  its  vital  functions  on  its  own 
substance,  as  animals  do  which  are  condemned  to 
starvation. 

The  ash  derived  from  water  in  which  yeast  has  been 
washed,  is  always  alkaline ;  that  of  the  washed  yeast 
is  acid;  which  induces  us  to  believe  that  there  is  an 
ammonio-magnesian  phosphate  present  in  the  yeast, 
which,  in  consequence  of  its  insolubiHty,  remains  in  the 
washed  ferment.  M.  Dubrunfaut  has,  like  M.  Pasteur, 
ascertained  the  constant  disappearance  of  a  certain 
portion  of  the  ammonia  of  the  medium,  as  a  concomitant 
of  the  reproduction  of  ferment ;  but  at  the  same  time, 
the  ash  becomes  more  acid.  The  addition  of  ammonia- 
cal  salts  to  fermentation  going  on  under  unfavourable 
conditions,  has  the  effect  of  facilitating  the  fermentation, 
and  preserving  the  proportion  of  nitrogen,  O'l  of  the 
yeast.* 

M.  Dubrunfaut  performed  a  series  of  experiments  to 
ascertain  the  part  played  by  different  mineral  salts  in 
the  fermentation  of  sugar,  and  in  the  development  of 
yeast.  He  made  use  for  this  purpose  of  solutions  of 
pure  sugar  at  lo  per  cent.,  and  added  to  them  various 
mineral  salts  and  yeast  in  the  form  of  paste,  all  taken 
at  a  weight  0*05  of  the  weight  of  the  sugar.  The  ferment 
employed,  therefore,  only  represented  in  dry  matter  o*oi 
of  the  weight  of  sugar.  He  tried,  I.  potassium  nitrate; 
2.  ammonium  sulphate  ;  3.  potassium  sulphate  ;  4.  cal- 
cium phosphate;  5.  magnesium  sulphate;  6.  calcium 

*  These  last  experiments  would  have  been  more  properly  placed  in  the 
chapter  concerning  the  influence  of  salts  and  nitrogenous  matter,  on  the  de- 
velopment of  yeast.  We  only  give  them  here  as  a  supplement,  which  the 
reader  will  be  kind  enough  to  read  for  himself  in  its  proper  place. 


3 


158 


ON  FERMENTATION. 


sulphate  ;  7.  sodium  sulphate  ;  8.  a  wort  without  mineral 
salts,  to  serve  for  comparison  : — 

FRACTIONS  OF  SUGAR  DECOM- 
POSED IN  THE  SAME  TIME, 


Sodium  sulphate  ....  0*52. 

Calcium      „  ....  0*62. 

Magnesium  „  ....  073. 

Calcium  phosphate  ....  o'8o. 

Potassium  sulphate  ....  o*88. 

Ammonium   „  .      .       •      .  0*94. 

Potassium  nitrate  .       .       .       ,  i*io 


for  complete  and  perfect  transformation  without  pro- 
duction of  acid. 

All  these  salts  gave  higher  results  than  those  of  the 
wort  used  for  comparison,  which  only  transformed  0*50 
of  the  sugar  into  alcohol. 

The  nitric  acid,  in  this  experiment,  entirely  dis- 
appeared. It  appears,  then,  from  Dubrunfaut's  observa- 
tions, that  yeast  forms  no  exception  with  respect  to  the 
facility  with  which  nitric  acid  is  assimilated. 


CHAPTER  VI. 


ACTION     OF     VARIOUS     CHEMICAL     AND  PHYSICAL 
AGENTS  ON  ALCOHOLIC  FERMENTATION. 

It  has  long  been  known  that  certain  chemical  com- 
pounds, especially  those  which  coagulate  albuminous 
substances  and  disorganize  the  tissues,  or  which,  by 
their  presence  in  sufficient  quantities,  are  incompatible 
with  life,  are  opposed  to  fermentation  ;  such  are  the 
acids  and  alkalies  in  suitable  proportions,  silver  nitrate, 
chlorine,  iodine,  the  soluble  iron,  copper  and  lead,  salts, 
tannin,  phenol,  creosote,  chloroform,  essence  of  mus- 
tard, alcohol  when  its  strength  is  above  20  per  cent, 
hydrocyanic  and  oxalic  acids,  even  in  very  small  quan- 
tities. 

An  excess  of  neutral  alkaline  salts  or  sugar  acts  in 
the  same  manner,  by  diminishing  in  the  interior  of  the 
cell  the  minimum  quantity  of  water  which  is  necessary 
for  the  manifestation  of  its  vital  activity. 

The  red  mercury  oxide,  calomel,  manganese  peroxide, 
the  alkaline  sulphites  and  sulphates,  the  essences  of 
turpentine  and  of  lemon,  &c.,  also  interfere  with,  and 
destroy,  alcoholic  fermentation. 

Phosphoric  and  arsenious  acids  are,  on  the  contrary, 
inactive.    We  owe  to  M.  Dumas  (Ann.  Chim.  Phys., 


ON  FERMENTATION. 


Sth  series,  vol.  3,  1874,  p.  81)  a  very  elaborate  work  on 
the  question  which  we  are  now  considering ;  we  will 
give  here  the  principal  results.  This  illustrious  chemist 
first  proved,  by  many  experiments,  that  the  fermentation 
of  sugar  under  the  influence  of  yeast  can  be  studied 
like  any  regular  phenomenon,  which,  when  subject  to 
determinate  disturbing  forces,  would  be  able  to  show 
their  influence  with  precision. 

Thus,  at  24^  C.  (about  76''  F.)  20  grammes  of  yeast 
decompose  i  gramme  of  sugar  (glucose)  in  twenty-three 
or  twenty-four  minutes,  (the  mean  of  several  trials 
agreeing  with  each  other). 

In  another  series  of  experiments  it  was  found  that 
100  grammes  of  yeast  decompose,  at  the  same  tempera- 
ture, I  gramme  of  glucose  in  twenty-four  minutes. 
Thus,  yeast  acts  on  glucose  with  the  same  rapidity, 
in  the  proportion  of  20  grammes  of  yeast  to  one  of 
glucose,  as  of  100  to  i. 

Experiments  made  on  the  same  yeast,  with  cane-sugar 
and  glucose  separately,  showed  that  the  destruction  of 
I  gramme  of  cane-sugar  by  40  grammes  of  yeast  had  a 
maximum  duration  of  thirty-four  minutes,  w^hile  one 
gramme  of  glucose  required  but  sixteen  or  seventeen 
minutes. 

If  we  mix  some  beer-yeast  with  water,  and  add  to 
similar  portions  of  the  liquid,  containing,  for  example, 
150  cubic  centimetres  of  water  and  10  grammes  of 
yeast,  quantities  of  sugar  represented  by  0*5  grammes, 
I  gramme,  2  grammes,  4  grammes,  wc  find  that  the 
time  necessary  for  the  destruction  of  the  sugar  is 
exactly  proportional  to  its  quantity ;  or,  in  other  words, 
that  under  identical  circumstances  the  duration  of  the 


ACTION  OF  CHEMICAL  AND  PHYSICAL  AGENTS.  l6l 

fermentation  is  proportional  to  the  quantity  of  sugar,  it 
being  understood  that  the  yeast  is  in  excess. 

By  taking,  then,  for  the  length  of  the  co-ordinates 
measured  along  the  axis  of  the  abscissae  the  quantities 
of  sugar,  and  for  the  co-ordinates  measured  along  axis 
of  the  ordinates,  the  number  of  minutes  necessary  for 
the  disappearance  of  the  sugar,  we  obtain  a  straight 
line. 

M.  Dumas  estimates  approximately,  according  to 
his  results,  that  to  decompose  i  gramme  (15*4  grains) 
of  sugar  in  one  hour,  it  requires  400  milliards 
(400,000,000,000)  cells,  supposing  them  all  to  be 
active. 

The  fact  discovered  by  Dumas,  that  beyond  a  certain 
limit  an  excess  of  yeast  does  not  promote  the  ferment- 
ation of  a  given  quantity  of  sugar,  is  extremely  im- 
portant. 

In  fact,  if  the  decomposition  of  sugar  is  a  biological 
phenomenon  which  takes  place  in  the  interior  of  each 
cell  into  which  the  sugar  penetrates  ;  if  it  ought  to  be 
attributed  to  the  nutrition  of  the  globules  of  yeast,  it 
seems  that  the  rapidity  of  the  decomposition  of  the 
same  quantity  of  sugar  ought  to  increase  indefinitely 
vv^ith  the  number  of  cells  which  are  in  action,  in  the 
same  manner  as  in  a  meal,  the  quantity  of  food  con- 
sumed increases  with  the  number  of  guests. 

Temperatitre. — We  have  to  determine  three  data 
relating  to  alcoholic  ferment,  with  respect  to  heat: — 

I.  What  are  the  two  limits,  maximum  and  minimum, 
compatible  with  the  life  of  the  ferment  1  2.  At  what 
temperature  is  alcoholic  fermentation,  that  is  to  say,  the 
activity  of  the  fernient,  the  greatest }  The  lower  limit 
8 


ON  FERMENTATION. 


compatible  with  the  hfe  of  the  Saccharomyces  is  not  yet 
determined  with  precision. 

It  is  only  towards  8°  or  ic<^  C.  (about  46°  to  50"^  F.) 
that  the  cell  shows  its  chemical  energy  by  sensible 
reactions  (fermentation,  absorption  of  oxygen,  &c.),  but 
it  may  be  cooled  with  impunity  towards  zero  (32^  R),  and 
even  below,  if  we  are  careful  to  subject  it,  after  con- 
gelation, only  to  very  slow  progressive  variations,  to 
re-melt  the  frozen  water  in  which  it  is  contained.  The 
higher  limit  depends  on  whether  we  operate  on  yeast 
previously  dried  or  still  damp. 

Thus,  yeast  dried  carefully  may  uc  heated  to  1 00*^ 
(212^  F.)  without  losing  its  vitality. 
1^    Damp  yeast  would  lose  its  activity  (Mayer)  towards 
/  53""  C.  (about  128"  F.),  or  at  least  at  a  temperature  less 
than  60^  C.  (140^  F.). 

As  to  the  most  favourable  conditions  of  temperature 
for  good  fermentation,  they  appear  to  range  between  25" 
and  30''  C.  (77''  and  86°  F.),  varying  within  narrow  Hmits, 
according  to  the  nature  of  the  medium  and  that  of  the 
ferment ;  even  near  zero  (32®  F.)  the  production  of 
alcohol  by  yeast  is  not  absolutely  destroyed. 

Electricity, — The  sparks  of  a  Holtz  machine,  or  those 
of  an  induction  coil  passing  through  water  containing 
yeast,  modify  neither  its  power  of  changing  cane-sugar 
into  glucose,  nor  its  activity  as  an  alcoholic  ferment. 

Gas-light. — The  process  of  fermentation  is  more  slow 
in  the  dark.  M.  Dumas  placed  becr-yeast  of  the  con- 
sistence of  thick  broth  in  flasks  respectively  filled  with 
oxygen,  hydrogen,  nitrogen,  carbon  oxide,  nitrogen  pro- 
toxide, proto-carburcttcd  hydrogen.  At  the  end  of  three 
days,  this  yeast,  placed  in  contact  with  sugar,  acted  in 


ACTION  OF  CHEMICAL  AND  PHYSICAL  AGENTS.  1 63 

the  same  way  as  yeast  kept  in  the  open  air,  and  the 
microscope  showed  no  change. 

AlcohoKc  fermentation  is  more  slow  in  a  vacuum. 

Siilplmr. — The  presence  of  flour  of  sulphur,  even  in 
equal  proportions  to  that  of  the  dried  yeast,  did  not 
sensibly  interfere  with  alcoholic  fermentation,  as  had 
been  predicted  ;  only  the  carbon  dioxide  which  was 
disengaged  contained  I  or  2  per  cent,  of  sulphuretted 
hydrogen. 

Action  of  Acids,  —  Yeast  is  always  acid.  If  we 
neutralize  the  free  acid  which  is  found  in  it  by  lime- 
water,  we  find  that  in  about  five  minutes  the  acidity 
reappears.  We  are  therefore  obliged,  in  order  to  main- 
tain a  continuous  neutrality,  to  add  for  each  gramme  of 
yeast  under  examination,  a  quantity  of  lime  equivalent 
to  '003  grammes  of  normal  sulphuric  acid.  M.  Dumas 
wished  to  ascertain  whether  the  acidity  of  yeast  can 
be  increased,  without  its  power  being  impaired,  and 
whether  the  specific  nature  of  the  acids  exercises  any 
influence  on  the  result. 

Experimenting  on  sulphuric,  sulphurous,  nitric,  phos- 
phoric, arsenous,  boracic,  acetic,  oxalic,  and  tartaric 
acids,  he  added  to  a  mixture  of  sugar,  water,  and  yeast 
(5  grammes  yeast,  10  grammes  sugar,  25  grammes 
water)  quantities  of  each  of  these  acids  respectively, 
first  equivalent,  then  tenfold,  and  a  hundredfold  of  the 
acid  power  of  the  yeast. 

The  addition  of  one  of  these  acids,  even  in  feeble 
proportions,  neither  hastened  the  commencement  nor 
the  end  of  the  fermentation.    It  often  arrested  the  de-  1 
composition  of  the  sugar.    In  general,  when  we  add  a  I 
quantity  of  acid  equal  to  100  times  the  weight  of  acid 


164 


ON  FERMENTATION. 


contained  in  the  yeast,  fermentation  does  not  take 
place.  It  was  necessary  to  add  200  equivalents  of 
hydrocloric  or  tartaric  acid  to  attain  this  result, 
although  ten  equivalents  of  these  acids  are  sufficient  to 
render  the  fermentation  slow  and  incomplete. 

Action  of  bases. — M.  Dumas  having  set  going  at  the 
same  time  eight  experiments  of  fermentation,  differing 
from  each  other  only  by  increasing  additions  of  ammonia 
from  O*  (quantities  equivalent  to  o,  I,  2,  3,  4,  8,  16,  24 
times  the  acid  of  the  yeast  employed),  observed  that  in 
the  first  five  vessels  the  fermentation  was  equal.  It  is 
only  in  proportions  equal  to  eight,  or  even  sixteen  times 
the  acid  of  the  yeast,  that  it  begins  to  proceed  more 
slowly  ;  but  at  the  end  of  some  hours  the  yeast  has 
again  become  acid.  In  the  flask  containing  a  quantity 
of  ammonia  equivalent  to  twenty-four  times  the  acidity 
of  the  yeast,  no  fermentation  takes  place. 

Yeast  seems  to  possess  the  power  of  producing  or 
exhaling  an  acid  which  neutralizes  the  bases  in  contact 
with  it ;  but  this  power  is  limited.  In  fact,  by  adding 
slaked  lime  or  calcined  magnesia  in  quantities  equal  to 
half  the  weight  of  the  yeast  there  will  be  no  fermenta- 
tion. The  oxides  of  zinc,  of  iron,  or  even  litharge,  have, 
on  the  contrary,  no  influence.  The  alkalies  tend  to  arrest 
fermentation,  but  do  not  destroy  it,  except  when  the 
proportion  is  rather  large.  Sodium  carbonate  has  no 
cff"ect,  unless  we  raise  the  proportion  to  70  grammes  for 
10  grammes  of  yeast,  and  200  cubic  centimetres  of 
sweetened  water  containing  yV  of  sugar.  Magnesium 
subcarbonate  has  no  action. 

Action  of  salts, — M.  Dumas  allowed  yeast  to  remain 
three  days  in  saturated  solutions  of  various  salts,  and 


ACTION  OF  CHEMICAL  AND  PHYSICAL  AGENTS.  165 

then  tried  its  action  on  the  solution  of  sugar-candy  ; 
his  experiments  led  him  to  class  salts  in  four  categories, 
viz : — 

1st.  Those  under  whose  influence  the  fermentation  of 
sugar  is  entire,  and  more  or  less  rapid — 


Potassium  sulphate 
„  chloride 
„  phosphate 
„  sulphovinate 
„  sulphomethylate 
„  hyposulphate 

Calcium  hyposulphate 

Potassium  formiate 
„  tartrate 
„  bitartrate 
„  sulphocyanide 
ferrocyanide 
ferricyanide 


Sodium  phosphate 
„  sulphate 
„  bisulphate 

pyrophosphate 
„  lactate- 

Ammonium  phosphate 

Magnesium  sulphate 

Calcium  chloride 
„  phosphate 
„  sulphate 

Strontium  chloride 

Alum 

Zinc  sulphate 

Cupric  sulphate  at  i^^gg 


2nd.  Those  under  whose  influence  there  is  partial 
fermentation  of  the  sugar,  more  or  less  retarded — 


Potassium  bisulphite 
„  nitrate 
„  butyrate 
„  iodide 
„  arseniate 

Sodium  sulphite 

„  hyposulphite 

Potassium  hyposulphite 


Borax 
White  soap 
Ammonium  nitrate 
„  tartrate 
Seignette  salt 
Barium  chloride 
Ferric  sulphate  37^5 
Magnesium  sulphate 


3rd.  Those  under  whose  influence  there  is  greater  or 
less  change  of  the  sugar,  without  fermentation  — 


i66 


ON  FERMENTATION. 


Potassium  nitrate 
„  chromate 
„  bichromate 

Sodium  nitrate 


Marine  salt 
Sodium  acetate 
Sal  ammoniac 
Mercuric  cyanide. 


4th.  Those  under  whose  influence  there  is  neither 
change  nor  fermentation — 


Thus  among  the  salts  studied,  there  are  some  which 
favour  fermentation,  up  to  a  certain  point,  or  at  least 
allow  it  to  run  its  entire  course  without  interruption  ; 
such  as  potassium  tartrate.  There  are  others  which 
retard  fermentation  and  render  it  incomplete,  the  action 
being  arrested,  when  the  liquor  still  contains  much 
changed  sugar  in  contact  with  it. 

There  are  some  which  do  not  allow  it  to  be  set  up, 
although  the  sugar  has  been  partially  changed. 

There  are  also  some,  which  not  only  do  not  allow 
fermentation  to  be  set  up,  but  which  even  oppose  the 
change  of  the  sugar. 

Strychnine  has  no  effect  on  the  properties  of  yeast. 


Potassium  acetate 
„  cyanide 


Sodium  monosulphide. 


CHAPTER  VII. 

CAN  NOTHING  BUT  ALCOHOLIC  YEAST  EXCITE 
ALCOHOLIC  FERMENTATION? 

We  must  now  enter  on  a  question  of  great  importance 
in  the  history  of  fermentation  in  general.  Admitting 
with  M.  Pasteur,  and  all  those  who  have  preceded  and 
followed  him  in  this  inquiry,  that  alcoholic  fermentation, 
and  the  other  phenomena  of  the  same  order,  such  as 
lactic,  butyric,  fermentations,  &c.,  are  palpable  manifesta- 
tions of  certain  physiological  functions  of  ferments,  or 
organisms  of  an  inferior  order,  we  may  ask,  whether 
the  power  of  resolving  glucose  into  alcohol  and  carbon 
dioxide,  or  of  changing  it  into  lactic  acid,  and  that 
again  into  a  mixture  of  hydrogen,  carbon  dioxide,  and 
butyric  acid,  belongs,  for  each  special  fermentation,  only 
to  a  single  organism,  to  a  single  ferment,  or,  at  least, 
to  species  very  nearly  allied,  as  we  have  seen  in  the 
species  of  the  genus  SaccJiaromyces ;  or  whether  these 
reactions  are  the  result  of  cell  life  in  general,  when 
organic  cells  are  placed  under  special  conditions.  On 
this  hypothesis,  ferments  would  have  no  advantage  over 
other  living  cells,  ,  than  that  of  showing  these  mani- 
festations in  a  more  energetic  and  intense  degree. 

It  is  easy  to  perceive,  a  priori,  what  new  horizons 
would  be  opened  for  biological  chemistry  by  such  an 


ON  FERMENTATION. 


idea,  if  it  were  correct,  and  what  a  vast  field  of  experi- 
ments would  be  offered  to  chemical  physiology. 

There  is  no  doubt  that  this  thought  must  have 
presented  itself  involuntarily,  as  we  may  say,  to  the 
minds  of  men  of  science  accustomed  to  reflect  on  these 
delicate  questions  of  the  reactions  in  living  organisms; 
but  M.  Pasteur  has  the  honour  of  first  clearly  expressing 
it,  and  supporting  it  by  positive  experiments.  (Pasteur, 
Comp.  Rend,  de  I'Acad.  des  Sciences,  vol,  75,  p.  784.) 
These  experiments  were  undertaken  in  order  to  prove 
that  the  alcoholic  fermentation  of  sugar  may  be  excited 
by  other  organisms  than  the  cells  of  Saccharomyces, 
and  notably  by  the  elementary  cells  of  larger  plants, 
such  as  are  found  in  fruits,  leaves,  &c.  The  researches 
of  M.  Lechartier  and  M.  Bellamy,  on  the  alcoholic  fer- 
mentation of  fruits  (Comp.  Rend,  de  TAcad.,  1869,  and 
1872,  vol.  75,  p.  1,203  ;  1874,  vol.  79,  p.  949,  and  1,006), 
have  been  directed  to  the  same  end,  and  lead,  as  we 
shall  see,  to  this  important  consequence,  that  the 
elementary  organs  of  plants  in  general  are  endowed, 
though  in  a  less  degree  than  the  cells  of  yeast,  with  the 
property  of  exciting  alcoholic  fermentation. 

Analogous  facts  had  been  observed  by  other  experi- 
mentalists ;  thus,  M.  Berard  informed  us,  as  early  as 
1 82 1,  that  when  fruits  are  placed  in  air,  or  in  oxygen 
gas,  a  certain  volume  of  this  gas  disappears,  whilst  at 
the  same  time  there  is  a  formation  of  a  nearly  equal 
volume  of  carbon  dioxide.  If  these  fruits  are  left,  on 
the  contrary,  in  carbon  dioxide  or  in  any  other  inert 
gas,  there  is  still  a  formation  of  carbon  dioxide  in  a 
notable  quantity,  "  as  if  by  a  sort  of  fermentation." 

M.  Fremy  had  also  shown  that  when  we  leave  barley 


ALCOHOLIC  FERMENTATION. 


ill  a  solution  of  sugar  in  water,  an  incontestable  intra- 
cellular fermentation  takes  place  in  the  interior  of  the 
corn.  (Comp.  Rend,  de  I'Acad.  des  Sciences,  vol.  75, 
p.  976  and  1,060.) 

The  same  investigator  observed  the  production  of 
alcohol  and  carbon  dioxide  in  the  interior  of  fruits, 
such  as  pears  and  cherries ;  but  guided  by  theoretical 
ideas  of  fermentation,  which  we  will  presently  explain, 
he  did  not  give  to  his  interesting  experiments  the 
interpretation  which  they  may  receive. 

M.  Fremy  admits  that  ferments  are  formed  under  the 
influence  of  the  living  organism.  Like  all  organisms 
in  process  of  development,"  said  he,  "  alcoholic  ferment 
may  present  itself  under  the  most  various  forms ;  it 
already  exists,  but  in  an  imperceptible  state,  in  the 
expressed  juice  of  the  grape,  which  appears  to  be 
transparent ;  it  soon  shows  itself  as  very  attenuated 
microscopical  corpuscles  ;  then  undergoing  a  fresh  de- 
velopment, it  is  precipitated  at  the  bottom  of  the  liquid, 
in  the  well-known  form  of  grains  of  yeast.  I  have 
frequently  examined  with  the  microscope  the  juice  and 
the  parenchyma  of  fruits,  before  or  after  their  ferment- 
ation, and  I  affirm  that  I  have  found  an  innumerable 
quantity  of  corpuscles  which  have  all  the  appearance 
of  organic  ferments.'* 

Thus  we  see,  from  this  extract,  that  the  intracellular 
fermentation  of  the  fruit  is  not  an  immediate  con- 
sequence of  the  biological  functions  of  the  living  cell ; 
there  is  an  intermediate  principle,  the  ferment,  produced 
by  the  organism,  which  excites  the  decomposition. 

He  ought,  in  a  historical  point  of  view,  to  mention 
that  the  experiments  of  M.  Lechartier,  and  M.  Bellamy, 


I/O 


QN  FERMENTATION. 


preceded  those  of  M.  Pasteur;  we  will  here  mention 
the  results  obtained  by  these  observers. 

First,  M,  Lechartier's  Researches,  —  This  observer 
placed  fruits  (pears,  apples,  lemons,  cherries,  chestnuts, 
medlars,  potatoes,  grains  of  wheat,  linseed,  currants)  in 
test-jars,  connected  with  smaller  test-tubes,  placed  over 
a  mercurial  trough.  Under  these  conditions,  the  whole 
of  the  oxygen  of  the  confined  air  in  which  the  fruits 
are  placed  is  absorbed.  This  absorption  is  accompanied 
and  followed  by  a  considerable  production  of  carbon 
dioxide  gas.  The  disengagement  of  carbon  dioxide  is 
generally  divided  into  two  distinct  periods ;  in  the  first, 
after  the  absorption  of  the  oxygen  gas  of  the  air  which 
remained  in  the  test-glasses,  the  disengagement  of 
carbon  dioxide  proceeds,  at  first,  in  a  regular  and 
uniform  manner,  then  it  slackens,  and  stops  for  a 
certain  time,  to  be  afterwards  renewed  with  an  in- 
creasing rapidity,  greater  than  that  observed  during  the 
first  period.  This  continues  for  several  months.  At 
this  time,  if  we  have  taken  the  precaution  to  experiment 
on  fruits,  isolated  from  each  other,  and  kept  from  con- 
tact with  the  sides  of  the  vessel  containing  them  ;  if, 
besides,  we  have  been  careful  to  prevent  any  deposit  of 
liquid  on  the  surface  of  the  fruit,  we  may  ascertain  the 
production  of  notable  quantities  of  alcohol,  easily 
separated  by  distillation,  after  the  fruit  that  has  been 
experimented  on  has  been  crushed  into  pulp  ;  and 
what  is  more,  a  careful  microscopical  examination  of  the 
parenchyma  reveals  no  trace  of  alcoholic  ferment. 

Thus,  on  November  12,  two  pears,  one  weighing  157 
grammes,  and  the  other  125,  were  suspended  sepa- 
rately, each  in  a  glass  jar,  well  corked,  and  provided 


ALCOHOLIC  FERMENTATION. 


171 


with  a  tube  by  which  gas  might  escape.  Calcium  chlo- 
ride had  been  previously  placed  at  the  bottom  of  the 
jars,  to  maintain  around  the  fruits  an  atmosphere  un- 
saturated with  the  vapour  of  water. 

The  test-jars  were  opened  on  July  19  :  1,762  cubic  - 
centimetres  of  gas,  and  2*62  grammes  of  alcohol  were 
collected.  The  pears  had  preserved  their  colour,  their 
skin  was  wrinkled,  but  not  damp.  Their  consistence 
and  smell  were  like  those  of  mellow  pears.  They  had 
lost  together  134  grammes  of  water,  yet  they  still  con- 
tained it  in  the  proportion  of  69  per  cent,  of  their 
weight. 

Microscopical  observations,  made  at  different  dis- 
tances from  the  centre,  could  discover  no  alcoholic 
ferment.  The  disengagement  of  carbon  dioxide  did 
not  take  place  in  a  regular  manner  ;  from  March  3  to 
April  8,  there  were  only  28  cubic  centimetres  evolved  ; 
and  from  April  8  to  July  19,  there  was  absolutely  none. 
The  existence  of  alcoholic  ferment  within  the  pear  is 
incompatible  with  all  cessation  of  activity  during  so 
considerable  an  interval  of  time.  This  fact,  therefore, 
corroborates  the  negative  results  obtained  by  means  of 
microscopical  examination. 

In  other  experiments,  arranged  in  the  same  manner, 
but  continued  much  longer,  M.  Lechartier  and  M.  Bel- 
lamy saw  the  disengagement  of  carbon  dioxide  resumed 
with  increasing  activity,  after  a  shorter  or  longer  period 
of  cessation.  In  these  cases,  we  can  clearly  ascertain 
the  presence  of  ferment.  We  may  suppose  that  the 
spores  and  germs  of  ferment,  which  exist  on  the  surface 
of  all  fruits,  as  now  seems  incontestable,  have  been  able 
to  penetrate  into  the  inside  of  the  fruit  by  means  of 


172 


ON  FERMENTATION, 


artificial  fissures,  and  meeting  there  an  appropriate 
medium,  have  budded  and  set  up  the  alcoholic  ferment- 
ation due  to  ferment 

The  different  fruits  mentioned  above  all  gave  ana- 
logous results,  and  the  experiment  before  described 
may  serve  as  the  type  of  the  rest,  which  were  very 
numerous,  and  were  published  by  the  authors  ;  we  will 
abstain  from  further  allusion  to  them  here. 

The  cessation  of  the  disengagement  of  carbon  dioxide 
may  last  a  long  time  ;  thus,  pears  have  been  preserved 
inert,  at  the  season  of  the  year  when  the  heat  is 
greatest,  during  a  time  varying  from  31  to  272  days; 
and  when  an  end  was  put  to  the  experiments,  there 
was  no  reason  to  suppose  that  this  state  of  things 
would  be  modified. 

After  this  internal  action,  the  fruits  had  undergone 
great  modifications. 

(i.)  As  soon  as  the  fruits  were  exposed  to  contact 
with  the  air,  they  become  brown  throughout  their  whole 
mass,  like  fruits  that  have  become  sleepy  "  or  rotten. 
The  leaves  assume  the  colour  and  aspect  of  dead 
leaves. 

(2.)  The  cellular  tissue  is  either  partly  or  completely 
disaggregated.  Thus  "  duchesse  pears  at  the  end  of 
a  year,  were  like  a  mass  of  syrup  covered  with  a  skin. 

(3.)  A  fruit  which  has  lost  its  activity  does  not  regain 
it,  even  after  being  placed  again  in  contact  with  the 
air.  • 

(4.)  The  germ  contained  in  the  fruit  participates  in 
the  decay,  and  the  seed  loses  its  property  of  germina- 
tion. 

It  appears  then,  as  these  writers  say,  that  at  the 


ALCOHOLIC  FERMENTATION. 


moment  when  fruits,  seeds,  and  leaves  are  detached 
from  the  plant  which  bears  them,  life  is  not  extinct  in 
the  cells  of  which  they  are  composed.  This  life  goes 
on,  sheltered  from  the  air,  consuming  sygar  and  pro- 
ducing alcohol  and  carbon  dioxide.  The  moment  when 
carbon  dioxide  ceases  to  be  formed,  is  that  at  which 
all  the  vitality  of  their  cells  is  destroyed.  Fruits,  seeds, 
and  leaves  may  then  remain  for  an  indefinite  period  in 
an  inert  state,  if  no  organic  ferment  is  developed  in  the 
interior. 

As  to  beet-root  and  potatoes,  an  especial  fact  has 
been  observed  which  ought  to  be  mentioned. 

The  whole  of  the  phenomena  are  the  same  as  with 
apples  and  pears ;  when  the  period  of  cessation  arrives, 
no  alcoholic  ferment  is  observed,  but  in  the  acid  liquid 
which  impregnates  the  mass  of  their  softened  or  dis- 
aggregated tissues,  we  find  bacteria  of  difi*erent  sizes, 
and  yet  we  can  detect  no  disengagement  of  carbon 
dioxide. 

Secondly,  M,  Pasteur  s  Experiments.  —  The  experi- 
ments made  by  M.  Pasteur,  while  they  verify  the  pre- 
ceding conclusions,  were  especially  intended  to  establish 
particular  views  respecting  fermentations,  and  a  general 
theory  of  these  biological  manifestations.  The  moment 
has  now  arrived  to  allude  to  them,  and  discuss  them. 
We  cannot  do  better  than  quote  the  exact  words  of  the 
author  (Pasteur,  Comp.  Rend,  de  TAcad.  des  Sciences, 
vol.  75,  p.  784,  et  seq.). 

I  have  been  inclined  for  a  long  time  to  consider 
fermentations,  properly  so  called,  as  chemical  pheno- 
mena, co-relative  with  physiological  actions  of  a  pecu- 
liar nature.    Not  only  have  I  shown  that  their  ferments 


174 


ON  FERMENTATION. 


are  not  dead,  albuminoid  matter,  but  actual  living 
organisms  ;  I  have  also  excited  the  fermentation  of 
sugar,  lactic  acid,  tartaric  acid,  glycerine,  and,  in  more 
general ,  terms,  of  all  fermentable  matters,  in  media 
exclusively  mineral,  an  incontestable  proof  that  the 
decomposition  of  fermentable  matter  is  correlative  with 
the  life  of  the  ferment,  that  it  is  one  of  its  essential 
aliments ;  for  instance,  under  the  conditions  to  which  I 
refer,  it  is  impossible,  that  in  the  constitution  of  the 
ferments  which  arise,  there  can  be  a  single  atom  of 
carbon  which  is  not  taken  from  the  fermentable  matter. 

That  which  distinguishes  the  chemical  phenomena 
of  fermentations  from  a  number  of  others,  and  par- 
ticularly from  the  functions  of  ordinary  life,  is  the  fact 
of  the  decomposition  of  a  weight  of  fermentable  matter 
much  greater  than  that  of  the  weight  of  ferment  in 
action. 

I  have  long  suspected  that  this  peculiar  character 
may  be  allied  to  that  of  nutrition  without  contact  of 
free  oxygen.  Ferments  must  be  living  organisms,  but 
of  a  peculiar  nature,  in  this  sense,  that  they  have  the 
property  of  exercising  all  the  functions  of  their  life,  not 
excepting  their  multiplication,*  without  necessarily  em- 
ploying the  oxygen  of  the  atmospheric  air. 

Let  us  call  to  mind  those  curious  infusoria  which 
cause  butyric  or  tartaric  fermentation,  or  certain  kinds 
of  putrefaction,  and  which  not  only  are  able  to  live  and 
multiply  without  contact  with  oxygen,  but  which  perish 
and  cease  to  excite  fermentation  if  we  dissolve  this  gas 
in  the  medium  in  which  they  feed.  This  is  not  all. 
By  careful  experiments,  made  with  the  yeast  of  beer, 

*  Not  iliat  of  sporulation.    Note  by  M.  Schutzenberger. 


ALCOHOLIC  FERMENTATION. 


I  have  shown,  that  if  the  hfe  of  this  ferment  were  carried 
on  partially  by  the  influence  of  free  oxygen  gas,  this 
little  cellular  plant  lost,  in  proportion  to  the  intensity 
of  this  influence,  a  part  of  its  fermenting  character,  that 
is  to  say,  that  the  weight  of  yeasty  which  is  proditced 
tender  these  conditions  duri7tg  the  decomposition  of  stigar^ 
increases  progressively^  and  approaches  the  weight  of 
decomposed  sugar ^  in  exact  proportion  as  its  life  goes  on 
the  presence  of  increasing  quantities  of  free  oxygen 
gas. 

"  Guided  by  all  these  facts,  I  have  been  gradually  led 
to  look  upon  fermentation  as  a  necessary  consequence 
of  the  manifestation  of  life,  when  that  life  takes  place 
without  the  direct  combustion  due  to  free  oxygen. 

"We  may  partially  see,  as  a  consequence  of  this 
theory,  that  every  being,  every  organ,  every  cell,  which 
lives,  or  continues  its  life,  without  making  use  of  atmos- 
pheric air,  or  which  uses  it  in  a  manner  insufficient  for 
the  whole  of  the  phenomena  of  its  own  nutrition,  must 
possess  the  characteristics  of  a  ferment  with  regard  to 
the  substance  which  is  the  source  of  its  total  or  com- 
plementary heat  It  appears  that  this  substance  must 
necessarily  be  oxygenated  and  carbonated,  since,  as  I 
stated  just  now,  it  serves  as  food  to  the  ferment.  I 
have  just  brought  to  the  support  of  this  new  theory, 
which  I  have  already  several  times  brought  forward, 
though  with  some  hesitation,  since  the  year  1861,  some 
new  facts  which  will  now,  I  hope,  cause  it  to  be  re- 
ceived." 

In  a  saccharine  liquid,  suited  to  the  nourishment  of 
ferments,  contained  in  a  vessel  which  allows  an  artificial 
sowing,  while  it  prevents  the  spontaneous  sowing  of 


176 


ON  FERMENTATION. 


aerial  germs,  M.  Pasteur  placed  on  the  surface  a  trace 
of  pure  Mycoderma  vmi,^  On  the  following  days  the 
growth  covers  by  degrees  all  the  liquid,  under  the  form 
of  a  continuous  film.  We  can  easily  ascertain  that, 
under  these  conditions,  the  development  of  the  Myco- 
derma gives  rise  to  an  absorption  of  atmospheric 
oxygen,  which  is  replaced  by  an  almost  equal  volume 
of  carbon  dioxide,  and  that,  on  the  other  hand,  no 
alcohol  at  all  is  formed. 

M.  Pasteur  having  already  pointed  out  that  the 
Mycoderma  vini  has  the  property,  when  it  grows  on  the 
surface  of  an  alcoholic  liquid,  of  burning  the  alcohol 
already  formed,  yielding  not  acetic  acid  or  aldehyde, 
but  water  and  carbon  dioxide,  it  appears  to  us  that  we 
cannot  conclude  from  the  absence  of  observed  alcohol 
in  the  experiment,  that  none  has  been  formed.  It  is 
quite  possible  that  it  may  have  undergone  a  complete 
combustion  at  the  very  moment  of  its  ehmination.  The 
volume  of  carbon  dioxide  disengaged  in  the  experiments 
was  nearly  equal  to  the  volume  of  oxygen  consumed  ; 
therefore,  unless  we  suppose  that  the  combustion  only 
acted  on  the  hydrocarbons,  or  on  carbon  itself,  we 
cannot  explain  this  equality ;  it  might  be  due  to  the 
disengagement  of  carbon  dioxide  under  the  influence 
of  an  alcoholic  fermentation,  one  of  the  characteristics 
of  which,  alcohol,  was  burnt  as  fast  as  it  was  produced. 
However  this  may  be,  let  us  return  to  M.  Pasteur*s 
experiment.    Let  it  be  repeated  exactly  under  the  same 

*  In  a  publication  previous  to  this,  M.  Pasteur  had  announced  that  the 
sedimentary  yeast  was  produced  by  the  mycoderma  vini,  deprived  of  contact 
with  oxygen;  M.  Pasteur  has  reconsidered  this  opinion,  which  he  no  longer 
thinks  thoroughly  well  founded. 


ALCOHOLIC  FERMENTATION. 


177 


conditions,  with  this  difference  only,  that  when  the  film 
is  complete,  the  vessel  be  shaken,  so  as  to  break  up 
this  film,  and  sink  it  as  far  as  possible,  for  the  fatty 
matter  which  accompanies  it  hinders  its  being  entirely 
wetted. 

On  the  next  day,  and  often  after  only  a  few  hours, 
when  the  experiment  is  made  at  a  temperature  of  from 
25°  to_30°  C.  (77°  to  86°  F.),  we  see  small  bubbles  of  gas 
continually  rising  from  the  bottom  of  the  vessel,  which 
show  that  fermentation  has  commenced  in  the  sac- 
charine liquid.  It  continues  during  the  following  days, 
although  very  feebly,  and  the  presence  of  a  sensible 
quantity  of  alcohol  in  the  liquid  is  readily  ascertained. 
A  careful  examination  of  the  cells,  or  joints  of  the 
submerged  Mycoderma,  by  means  of  the  microscope, 
shows  that  these  joints  do  not  reproduce. 

This  latter  fact  seems  to  disagree  with  the  opinion 
so  many  times  advanced  elsewhere  by  the  writer,  that 
alcoholic  fermentation  is  always  accompanied  by  the 
development  and  multiplication  of  ferment,  and  that 
it  is  a  consequence  of  the  formation  of  buds. 

The  experiment  which  we  have  just  described  in 
detail,  quoting  almost  literally  the  words  of  the  author, 
allows  us  to  conclude  with  certainty  that  the  Mycodenna 
villi  may  play  the  part  of  alcoholic  ferment,  when  it 
is  placed  in  contact  with  a  saccharine  medium  ;  but  we 
cannot  admit  the  other  consequences  that  M.  Pasteur 
tries  to  draw  from  it,  as  proofs  of  the  theory  before 
cited. 

We  may  say  the  same  thing  of  the  experiments  on 
fruits  placed  entirely,  as  M.  Pasteur  has  done,  in  an 
atmosphere  of  carbon  dioxide,  or  of  those  of  M.  Le- 


178 


ON  FERMENTATION. 


chartier,  in  which  this  condition  was  rapidly  obtained 
by  the  absorption  of  the  oxygen  of  the  surrounding 
air. 

They  prove  irrefragably  that  vegetable  cells  can  pro- 
duce alcohol  and  carbon  dioxide  at  the  expense  of 
sugar,  and  can  act  like  the  yeast  of  beer,  but  much  less 
energetically ;  but  nothing  in  the  description  given  of 
their  experiments  by  M.  Pasteur  and  M.  Lechartier 
can  induce  the  reader  to  admit,  that  this  production 
of  alcohol,  this  cellular  fermentation,  only  commences 
from  the  very  moment  when  the  cell  is  either  partially 
or  entirely  removed  from  contact  with  oxygen. 

Even  admitting,  which  the  writers  quoted  do  not 
assert,  that  they  have  directly  ascertained  the  absence 
of  alcohol  in  the  fruit,  as  long  as  it  was  not  kept  from 
the  influence  of  oxygen,  I  still  have  the  idea  that  the 
alcohol  formed  may  have  been  burned,  and  may  have 
disappeared  under  the  form  of  water  and  carbon 
dioxide.  Besides  this  the  yeast  of  beer,  the  Saccharo- 
myces  cerevisicB  shows  us  in  th-e  most  striking  and 
incontestable  manner,  an  instance  of  a  cell  which 
excites  alcoholic  fermentation,  even  in  the  presence  of 
free  oxygen  dissolved  in  the  medium  ;  and  now  that  it 
is  no  longer  recognized  as  the  sole  producer  of  alcoholic 
fermentation,  I  do  not  see  why  we  should  wish  to 
preserve  for  it  so  exceptional  a  character  as  that  of 
being  the  only  exciter  of  it  in  the  presence  of  oxygen. 
As  to  this  latter  fact,  we  have  only  to  quote  M.  Pasteur 
himself,  in  order  to  prove  its  reality  (Bullet.,  Soc.  Chi- 
miquc,  i86i,  p.  621).  "Yeast  already  formed  may  bud 
and  develop  itself  in  a  saccharine  and  albuminous  liquid 
in  the  entire  absence  of  air  or  of  oxygen.    Little  fcr- 


ALCOHOLIC  FERMENTATION. 


179 


ment  is  formed,  in  this  case,  and  a  comparatively  large 
quantity  of  sugar  disappears,  sixty  or  eighty  parts  for 
one  of  the  ferment  formed.  Fermentation  is  very  slow 
ztnder  these  conditions.  If  the  experiment  be  made  in 
contact  with  air,  and  over  a  large  surface,  the  fer- 
mejitation  is  rapid.  For  the  same  quantity  of  sugar  that 
disappears,  much  more  yeast  is  produced.  This  is 
developed  under  these  conditions  with  energy,  but  its 
character  as  a  ferment  tends  to  disappear.  We  find,  in 
fact,  that  for  one  part  of  ferment  formed,  there  will  be 
only  from  four  to  ten  parts  of  sugar  transformed.  The 
part  of  ferment  played  by  this  yeast,  nevertheless, 
remains,  and  shows  itself  very  energetic^  if  we  cause  it 
to  act  on  sugar  when  not  under  the  influence  of  free 
oxygen  gas. 

M.  Pasteur  explains  the  fact  of  a  tumultuous  activity 
at  the  commencement  of  the  fermentation,  by  the 
influence  of  the  oxygen  of  the  air  which  is  dissolved  in 
the  liquids  when  the  action  commences. 

There  seems  to  be  some  contradiction  between  the 
conclusions  and  the  facts.  On  one  hand,  it  is  ascertained 
that  fermentation  in  the  presence  of  free  oxygen  is 
very  rapid  and  tumultuous  ;  on  the  other  hand,  it  is 
concluded  that  yeast  loses  its  character  as  a  ferment  in 
the  presence  of  oxygen.  In  reality,  this  contradiction 
disappears  if  we  measure,  as  M.  Pasteur  has  done,  the 
power  of  the  ferment,  not  by  the  quantity  of  sugar 
decomposed  by  the  unit  of  weight  of  yeast,  in  the  unit 
of  time,  which  would  lead  us  to  consequences  the  re- 
verse of  those  of  this  observer,  but  by  the  ratio  between 
the  weight  of  the  sugar  decomposed,  and  that  of  the 
ferment  formed  by  budding.    According  to  M.  Pasteur, 


i8o 


ON  FERMENTATION. 


fermentation  is  a  consequence  of  budding,  and  one  of 
the  phenomena  may  form  the  measure  of  the  other. 

Thus,  by  ascertaining  that  in  fermentation  without 
oxygen,  the  ratio  between  the  sugar  decomposed,  and 
the  yeast  formed,  is  from  60  or  80  to  i,  while  in  fer- 
mentation, in  presence  of  oxygen,  it  is  only  4  or  10  to 
I,  M.  Pasteur  thinks  himself  justified  in  concluding 
that,  although  in  this  latter  case  the  fermentation  is 
more  active,  the  character  as  a  ferment  of  the  yeast  has 
been  lowered. 

We  may  first  remark  that,  in  order  to  measure  the 
energy  of  a  ferment,  M.  Pasteur  makes  use,  on  this 
occasion,  of  an  hypothesis  which  admits  of  discussion. 

Is  it  not  more  natural  to  measure  the  energy  of  a 
ferment  by  the  effect  which  it  produces,  and,  in  the 
present  case,  by  the  quantity  of  sugar  decomposed  in 
the  unit  of  time  by  the  unit  of  weight  ? 

That  which  does  come  out  clearly  from  the  interest- 
ing facts  observed  by  M.  Pasteur,  is,  that  yeast  is  able 
to  multiply  in  ft  suitable  albuminous  saccharine  medium, 
with  or  without  the  presence  of  free  oxygen ;  the 
mutiplication  is  greatly  favoured  by  oxygen,  which 
explains  why  the  ratio  between  the  sugar  split  up  and 
the  yeast  formed  passes  from  80.1,  to  4.1,  but  at  the 
same  time  the  activity  of  the  ferment  is  notably  in- 
creased, and  the  fermentation  is  rapid  and  tumultuous. 

The  following  experiment  is  also,  in  my  opinion, 
opposed  to  M.  Pasteur's  theory.  I  have  carefully 
studied,  by  means  of  a  rapid  and  exact  method  of 
quantitative  analyses  of  the  dissolved  oxygen,  which 
has  been  described  elsewhere,  the  respiratory  pheno- 
mena of  yeast.    I  have  thus  been  able  to  ascertain  tliat 


ALCOHOLIC  FERMENTATION.  l8l 

in  a  liquid  medium  containing  oxygen  in  solution,  the 
quantity  of  oxygen  absorbed  by  one  gramme  of  yeast, 
in  the  unit  of  time,  and  at  the  same  temperature,  was 
constant,  whether  the  liquid  be  saturated  or  super- 
saturated with  oxygen,  or  contain  a  quantity  of  oxygen 
less  than  is  required  for  saturation.  Thus,  by  diffusing 
one  or  two  grammes  of  yeast  in  a  litre  of  aerated  water, 
we  notice  a  sensible  decrease  of  the  respiratory  power, 
only  when  the  proportion  of  oxygen  is  less  than  one 
cubic  centimetre  per  litre.  On  the  other  hand,  these 
respiratory  phenomena  follow  the  same  law  in  a  sac- 
charine and  oxygenated  medium  containing  nutritive 
albuminoid  matter,  but  the  intensity  of  the  respiration 
is  increased  ;  that  is  to  say,  that  the  proportion  of 
oxygen  absorbed  in  the  unit  of  time,  by  the  unit  of 
weight,  is  more  considerable  at  the  same  temperature 
than  in  pure  water.  Finally,  I  have  measured  the 
alcohol  produced,  in  a  given  time,  in  two  fermentations 
placed  identically  under  the  same  conditions,  and  with 
this  difference  only,  that  in  the  one  oxygen  was  con- 
tinually kept  dissolved.  The  proportion  of  alcohol 
was  found,  in  this  case,  to  be  sensibly  greater,  which 
agrees  with  Pasteur's  experiments ;  during  the  whole 
duration  of  the  fermentation,  we  have  been  able  to 
detect  a  rapid  absorption  of  oxygen,  the  intensity  of 
which  corresponded  with  the  general  results  obtained  in 
many  experiments. 

If,  as  M.  Pasteur  asserts,  the  decomposition  of  sugar 
were  the  result  of  a  respiration  of  the  cells  of  yeast  at 
the  expense  of  the  combined  oxygen,  recruiting  the 
free  oxygen,  it  seems  evident  that  fermentation  ought 
not  to  take  place,  or  at  least  ought  to  be  sensibly 


1 82  ON  FERMENTATION. 

lessened,  in  the  presence  of  oxygen  ;  but  the  reverse 
of  this  is  the  case.  The  respiratory  power  of  yeast  is 
independent  of  the  quantity  of  oxygen  contained  in  the 
medium  in  which  it  hves ;  it  only  varies  with  the  tem- 
perature, and  the  more  or  less  favourable  conditions  of 
nutrition,  as  well  as  with  the  more  or  less  perfect  state 
of  health  of  the  cell. 

Yeast  placed  in  a  saccharine  and  oxygenated  medium 
breathes  as  actively,  or  even  more  so,  than  in  pure 
water ;  it  therefore  fully  satisfies  its  respiratory  power 
at  the  expense  of  free  oxygen,  and  yet  it  excites  the 
fermentation  of  the  sugar  as  actively,  or  even  more  so, 
than  in  a  medium  which  is  not  oxygenated.  The  con- 
clusion to  be  drawn  from  this  is  plain  ;  the  respiratory 
power,  and  the  fermenting  power,  are  two  qualities 
inherent  in  the  cell  of  SaccharomyceSy  but  independent 
of  each  other,  in  this  sense,  that  one  of  these  phe- 
nomena is  not  the  consequence  of  the  other.  The 
energy  of  these  two  powers  depends  on  conditions  more 
or  less  favourable  to  the  nutrition  and  development  of 
the  yeast ;  it  is  not,  therefore,  surprising  to  see  them 
increase  or  grow  less  together.  In  other  words,  they 
are  not,  as  M.  Pasteur  supposes,  the  two  variable  terms 
of  a  constant  sum,  of  which  the  one  vanishes  when  the 
other  attains  its  maximum  value ;  on  the  contrary,  all 
the  facts  tend  to  prove  that  these  two  values  grow 
weak,  are  destroyed,  or  attain  their  maxima  at  the  same 
time,  under  the  influence  of  the  same  causes.  By 
weakening  the  vitality  of  the  yeast,  we  diminish  at  the 
same  time  its  respiratory  power,  and  its  power  as  a 
ferment ;  on  the  contrary,  by  placing  the  yeast  under 
the  best  physical  and  chemical  conditions  of  nutrition, 


ALCOHOLIC  FERMENTATION.  183 

Ave  see  the  two  factors  increase  side  by  side,  and  attain 
their  maximum  together.  Let  us,  however,  hasten  to 
add,  that  these  two  phenomena  are  necessarily  depen- 
dent on  each  other ;  and  we  cannot  deny  this  without 
contradicting  that  which  we  have  just  said. 

In  fact,  sugar  is  a  pabulum  necessary  to  the  develop- 
ment of  the  cells  of  the  Saccharomyces ;  its  presence  is, 
therefore,  indispensable  to  the  nutrition  of  these  cells. 
To  deprive  the  yeast  of  sugar,  or  which  amounts  to  the 
same  thing,  to  place  it  so  that  it  is  impossible  for  it 
to  act  as  a  ferment,  is  to  diminish  its  vitality,  and  to 
place  it  in  a  state  comparable  to  starvation  ;  therefore 
it  is  not  surprising  to  see  its  respiratory  power  diminish, 
all  other  things  being  equal.  Reciprocally,  to  prevent 
yeast  from  breathing  free  oxygen,  is  to  deprive  it 
of  one  of  its  most  important  functions,  and  again  to 
place  it  under  pathological  conditions  ;  and  its  activity 
as  a  ferment  will  be  lessened,  as  M.  Pasteur  himself 
observes. 

The  following  experiments  all  support  this  view. 
We  know,  by  the  researches  of  M.  Bechamp,  that  the 
yeast  of  beer  placed  under  conditions  of  starvation, 
that  is  to  say,  kept  from  contact  with  a  saccharine 
liquid,  undergoes  a  great  modification,  which  has 
nothing  in  common  with  putrefaction.  It  grows  soft, 
and  converts  its  protoplasm,  and  a  part  of  its  contents, 
into  soluble  principles,  among  which  M.  Bechamp  dis- 
tinguished leucine  and  tyrosine,  a  soluble  albuminous 
substance  coagulable  by  heat,  a  ferment  producing 
change,  a  gummy  substance,  phosphates,  and  acetic 
acid ;  these  phenomena  are  further  accompanied  by 
the  production  of  alcohol  and  carbon  dioxide,  as  well 


184 


ON  FERMENTATION. 


as  by  a  disengagement  of  pure  nitrogen.  I  have  myself 
closely  examined  this  singular  reaction,  and  have  been 
able  to  show,  in  the  extract  of  softened  yeast,  the  pre- 
sence of  xanthine,  hypoxanthine,  carnine,  and  guanine. 

After  these  first  researches,  I  thought  it  would  be 
interesting  to  study  the  modifications  which  took  place 
in  the  yeast  when  thus  changed  as  to  its  respiratory 
functions.  For  this  purpose  I  made  use  of  the  methods 
described  elsewhere,  when  treating  of  the  respiratory 
intensity  of  yeast.  A  known  weight  of  yeast  is  diffused 
in  a  known  volume  of  aerated  water,  the  original  oxy- 
metrical  degree  of  which  is  known;  this  degree  is  again 
measured  after  a  quarter  of  an  hour,  or  half  an  hour's 
respiration.    The  following  facts  were  observed. 

Four  hundred  grammes  of  fresh  yeast,  containing 
28-94  per  cent,  of  solid  fixed  matter,  were  diffused  in 
distilled  water,  so  as  to  former  2  litres  (about  3^  pints). 
This  homogeneous  liquid  was  divided  into  ten  equal 
parts,  and  introduced  into  bottles  nearly  filled.  These 
bottles  were  left  to  themselves  in  a  stove  heated  to 
20°  C.  (68°  F.). 

I  did  not  consider  it  to  be  necessary  to  employ  in 
these  experiments  the  creosote  water  recommended  by 
M.  Bechamp,  as  I  was  unacquainted  with  the  nature  of 
its  influence  on  oxymetrical  analyses  or  on  the  respira- 
tory power.  I  therefore  ran  the  risk  of  seeing  some 
vegetable  productions  make  their  appearance,  which 
were  foreign  to  the  yeast,  but  the  small  proportion  of 
which,  when  compared  with  the  yeast,  could  not  disturb 
the  results  by  any  very  large  error. 

At  the  end  of  three  days  without  nourishment,  the 
first  and  second  vessels  were  emptied  on  a  filter  which 


ALCOHOLIC  FERMENTATION. 


185 


had  been  previously  weighed  ;  that  which  remained  on 
the  filter,  was,  in  this  case,  well  washed  with  water  at 
30°  C.  (86''  F.),  until  it  was  exhausted,  then  dried  and 
weighed.  The  weight  of  the  filter  being  deducted, 
there  were  found,  for  the  40  grammes  of  original  yeast 
5*37  grammes,  or  13*4  per  cent,  of  solid  fixed  matter. 
The  contents  of  the  second  bottle  were  boiled  in  water 
before  filtration ;  the  fixed  residuum  found  was,  573 
grammes,  or  14  per  cent. 

After  digestion  for  six  days,  followed  by  boiling,  the 
third  bottle  gave  a  dry  fixed  residue  of  4*16  grammes, 
or  10*4  per  cent. 

After  a  longer  time,  the  diminution  in  weight  of  the 
solid  insoluble  materials  of  the  yeast  grew  less  by 
degrees,  and  became  insensible.  Fresh  yeast,  washed 
with  lukewarm  water,  loses  about  9*5  grammes  per  cent, 
of  soluble  matter.  Thus,  in  five  days,  the  yeast  which 
had  been  kept  without  nourishment,  transformed  into 
soluble  products,  i8'5— 9'5z=9*  grammes  of  insoluble 
matter,  which  resisted  the  first  washing. 

When  it'  has  reached  this  limit,  and  has  been  well 
washed  with  warm  water  till  it  runs  off  pure,  yeast 
shows  itself  completely  inactive  with  regard  to  oxygen. 
Thus  fresh  yeast,  at  20°  C.  (68°  F.),  respired,  per 
gramme  and  per  hour,  1*4  cubic  centimetre,  whilst  after 
exhaustion,  by  washing  the  absorption  of  oxygen,  under 
the  same  conditions,  did  not  exceed  "23  cubic  centi- 
metre. Yeast  digested,  without  nourishment,  for  forty- 
eight  hours  and  well  washed,  being  placed  in  a  pure 
saccharine  medium,  only  excited  a  very  slow  and 
scarcely  perceptible  fermentation,  as  M.  Bechamp  had 
before  observed. 

9 


ON  FERMENTATION. 


Thus,  digestion  without  nourishment,  when  sufficiently 
prolonged,  and  followed  by  thorough  washing,  affords 
us  a  very  simple  method  of  removing,  almost  entirely  and 
simultaneously,  the  two  most  characteristic  biological 
manifestations  of  the  yeast-cell.  Yet  its  vitality  is  not 
destroyed,  if  the  experiment  has  not  been  carried  too  far. 

If  we  add  to  aerated  water,  containing  yeast  which 
has  been  digested  and  washed,  and  which  only  acts  on 
oxygen  in  an  almost  imperceptible  manner,  some  water 
with  which  digested  yeast  has  been  washed,  we  shall  see 
the  respiratory  phenomenon,  at  first  very  feeble,  grow 
more  and  more  decided,  and  resume  an  activity  equal, 
and  even  superior,  to  that  of  fresh  yeast  simply  diffused 
in  pure  water.  The  fresh  yeast  itself  respires  more 
energetically  if,  instead  of  pure  water,  we  employ 
aerated  water  containing  soluble  products,  derived  from 
the  digestion  of  yeast. 

On  the  other  hand,  if  we  add  the  soluble  products  of 
yeast  to  the  mixture  of  digested  and  washed  yeast  and 
saccharine  water,  a  mixture  which  gives  rise  only  to  an 
insensible  fermentation,  we  see,  in  a  very  short  time,  the 
decomposition  of  the  sugar  manifest  itself  by  a  more 
abundant  disengagement  of  carbon  dioxide. 

It  follows,  from  the  whole  of  the  facts  noticed  in  this 
chapter,  that  we  can  no  longer  regard  alcoholic  fer- 
mentation as  the  simple  result  of  the  biological  inter- 
vention of  a  special  living  organism,  characterized  by  its 
form  and  by  the  conditions  of  its  development,  which 
alone  has  the  power  to  split  up  some  one  particular 
organic  compound  in  some  one  particular  way. 

The  various  kinds  of  fermentation  appear  to  us,  more 
and  more,  as  particular  cases  of  tlie  cliemical  activit}^  of 


ALCOHOLIC  FERMENTATION. 


living  cells,  and  the  ideas  enunciated  at  the  beginning  of 
this  work  on  fermentation  find  a  complete  confirmation 
in  the  new  facts  studied  in  this  chapter. 

M.  A.  Fitz  (Soc.  Chim.  de  Berlin,  January  and  Feb- 
ruary, 1873),  has  published  a  long  memoir  on  alcoholic 
fermentation  produced  by  the  Mucor  racemostcs. 

The  ratio  of  the  alcohol  to  the  carbon  dioxide  was 
found,  as  the  mean  of  many  determinations,  to  be  equal 
to   ^       while  in  the  alcoholic  fermentation  set  up  by 


Distilled  alcohol  contains  a  little  aldehyde ;  the 
residuum  contains  succinic  acid  ;  while  the  presence  of 
glycerine  is  doubtful.  Dextrin,  inulin,  and  sugar  of 
milk  do  not  ferment,  alcoholically,  in  the  presence  of 
Mticor  racemosics. 

With  altered  sugar,  the  fermentation  set  up  by  Mucor 
raceinostis  stops  after  a  short  time,  so  that  half  of  the 
sugar  remains  intact. 

The  cause  of  this  rapid  check  in  the  process  of  fer- 
mentation is  merely  the  sensitiveness  of  this  ferment  to 
the  influence  of  alcohol. 

M.  Fitz  has,  in  fact,  shown 'that  fermentation  ceases 
when  the  proportion  of  alcohol  in  the  liquid  reaches 
3*5  to  4  per  cent,  by  weight.  In  all  other  respects  the 
Mticor  raceinostis  behaves  like  yeast ;  it  alters  sugar, 
assimilates  the  nitrogen  of  ammoniacal  salts,  and  re- 
produces at  the  expense  of  sugar  (1*3  per  cent,  of  the 
sugar  decomposed). 

M.  Traube  (Soc.  Chim.  de  Berlin,  vol.  vii,  p.  887, 
1874)  has  studied  the  action  of  media  containing  no 
free  oxygen  on  alcoholic  ferments.  Hj^^SSpSriiOCi^, 


yeast  of  beer,  it  is  xoo. 

'  963 


lead  to. the  following  conclusions  : — 


i88 


ON  FERMENTATION. 


The  cell  of  the  ferment  is  not  developed  in  the 
absence  of  free  oxygen,  even  in  its  most  favourable 
medium,  the  must  of  grapes. 

The  ferment,  in  process  of  development,  continues 
to  increase  in  suitable  media,  even  in  the  absence  of  all 
trace  of  oxygen,  as  M.  Pasteur  had  already  shown- 
The  contrary  assertions  of  Brefeld  are  erroneous. 

M.  Pasteur's  theory,  according  to  which  the  yeast, 
in  the  absence  of  air,  takes  from  the  sugar  the  oxygen 
necessary  for  its  development,. is  not  well  founded;  in 
fact,  this  development  stops  long  before  the  greater 
part  of  the  sugar  is  decomposed.  Is  it  from  the  albu- 
minoid matter  that  the  ferment  takes  this  oxygen  in 
the  absence  of  air  ? 

Yeast  sets  up  alcoholic  fermentation  in  a  solution  of 
pure  sugar  in  the  absence  of  all  trace  of  oxygen,  but 
without  developing.  This  is  contrary  to  the  affirmation 
of  M.  Pasteur  that  fermentation  is  bound  up  with  the 
organization  of  the  yeast,  or  is  a  phenomenon  correla- 
tive to  the  vital  activity  of  the  cells. 


CHAPTER  VIIL 


VISCOUS  OR  MANNITIC  FERMENTATION  OF  SUGAR. 

Like  all  the  phenomena  v/hich  are  spontaneously 
produced  under  the  ordinary  conditions  of  humanlife, 
viscous  fermentation  has  been  known  for  a  long  time, 
for  it  is  developed  in  certain  wines  and  natural  saccharine 
juices,  as  the  juice  of  beetroot,  carrots,  and  onions,  as 
well  as  in  certain  juleps  and  potions  containing  sugar 
and  nitrogenous  matter,  and  it  becomes  apparent  by  the 
viscosity  of  the  liquid. 

This  reaction  was  studied  by  Braconnot  (Ann.  de 
Chim.  (i),  Ixxxvi.  p.  97,  1813),  Desfosse  (Journ.  de 
Pharm.,  xv.  p.  604),  Frangois  (Ann.  de  Chim.  et  de  Phys. 
(2),  vol.  xlvi.  p.  212,  1829),  Pelouze  and  Jules  Gay-Lussac 
(Ann.  de  Chim.  et  de  Phys.  (2),  vol.  Hi.  p.  410,  1833), 
Kircher  (Annalen.  Ch.  Pharm.,  xxxi.  p.  337),  Tilley  and 
Maclagan  (Philos.  Mag.,  xxviii.  p.  12),  Boutron-Charlard 
and  Fremy  (Comp.  Rend.,  xii.  p.  708,  1841),  and  also 
by  Pasteur  (Bullet,  Soc.  Chim.,  p.  30,  Paris,  1861). 

Viscous  fermentatation  is  excited,  according  to 
Pasteur,  by  a  special  ferment  acting  on  glucose  or  cane- 
sugar,  previously  altered,  transforming  them  into  a  kind 
of  gum  or  dextrin,  mannite,  and  carbon  dioxide.  Gum, 
mannite,  and  carbon  dioxide  are  the  only  constant  pro- 
ducts of  this  reaction  ;  the  lactic  acid,  butyric  acid,  and 


1 90 


ON  FERMENTATION. 


Iiydrogen,  which  we  see  appear  at  the  same  time,  are 
the  products  of  other  fermentations  due  to  foreign  fer- 
ments. 

M.  PeHgot  (Traite  de  Chimie  de  Dumas,  vol.  vi. 
P-  335>  1843)  was  the  first  to  notice  a  special  ferment, 
capable  of  producing  viscous  fermentation  in  saccharine 
solutions  to  which  it  is  added.  Pasteur  observed,  after- 
wards, that  this  ferment  is  formed  of  small  globules 
united  as  in  a  necklace,  whose  diameter  varies  from 
•0012  millimetres  (-000047  i^-)  '0014  ('000055  in.)  (fig. 
20).  These  globules,  sown  in  a  saccharine  liquid  contain- 


ing nutritive  nitrogenous  matter  and  mineral  substances, 
always  give  rise  to  viscous  fermentation:  100  parts  of 
sugar  give  about  51*09  of  mannlte  and  45*5  of  gum; 
besides  these,  carbon  dioxide  is  disengaged.  These 
results  may  be  shown  by  the  following  equation  : — 

No.  I.  25  (O2       Qii)  4-  25  (H2  O)  =  12  0^<>)  -f 

Cane-Sugar.  Water.  Gum. 

24  (a  W  O')  +  12  (C02)  +  12  (H2  0) 


Fig.  20.— Viscous  Ferment  of  Wine. 


Mannitc. 


Carb.  diox. 


V/ater. 


This  gives,  for  100  parts  of  sugar, — 


Mannlte 
Gum 

Carbon  dioxide 


51-09 
45-X8 
GIB 


MANNITIC  FERMENTATION  OF  SUGAR.  Ujl 

M.  Monoyer  (Th^se  pour  le  Doctorat  en  Medicine, 
Strasbourg,  1862)  proposes  to  represent  the  produc- 
tion of  mannite  and  of  gum  by  two  separate  equations, 
applying  to  two  distinct  phenomena,  independent  of 
each  other. 

No.  2.  13  (O2       0^2^)^  +  12  H2  O  =  24  (C^  W  O')  +  12  CO2 

No.  3.  12  {O-^  U^'  O^^)^  ==  12        H^o  O^^)  +24  H2  O  ; 

which,  added  together,  reproduce  Equation  No.  I  (plus 
12  molecules  of  water).  The  respective  proportions 
of  gum  and  mannite,  corresponding  to  the  preceding 
results,  are  obtained  generally  and  with  constancy  when 
we  sow  the  ferment  before  described. 

In  certain  viscous  fermentations,  however,  the  pro- 
portion of  gum  is  greater  than  that  of  mannite.  In 
this  case,  according  to  M.  Pasteur,  the  presence  in  the 
liquid  of  larger  globules  of  a  different  kind  is  perceived. 
The  same  writer  adds,  that  it  may  be  possible  that  this 
second  ferment  transforms  the  sugar  into  gum  only, 
without  there  being  any  ferment  ation  of  mannite.  To 
prove  the  accuracy  of  this  view,  it  would  be  necessary 
to  be  able  to  isolate  the  second  ferment  from  the  first, 
and  to  cause  it  to  act  separately :  hitherto  this  separa- 
tion has  not  been  effected.  However  this  may  be,  this 
variability  in  the  proportion  of  the  gum  tells  in  favour 
of  M.  Monoyer  s  opinion. 

The  liquids  which  are  most  apt  to  produce  viscous 
fermentation  -|-  can  also  undergo  lactic  and  butyric  fer- 

*  Perhaps  better  written,  13  (€«  H12  +  Ce  H>2  O^)  and  12  (C®  H>2  O^  + 
(^  6  JI12  0<^)  as  these  formulce  represent  the  "  changed  "  or  "  altered  "  cane- 
sugar,  i.e.,  cane-sugar  that  has  taken  up  one  molecule  of  water  and  split  up 
into  a  mixture  of  dextrose  and  levulose. 

t  Decoction  of  the  yeast  of  beer  filtered,  and  with  the  addition  of  sugar  : 


ig2 


ON  FERMENTATION. 


mentation  ;  but,  in  this  case,  the  organized  forms  of  hfe 
which  are  developed  in  the  hquid  are  of  a  different 
nature. 

The  gum  which  is  formed  under  these  conditions  is 
more  alHed,  by  its  characters,  to  dextrine  than  to  gum 
arabic.  Nitric  acid  converts  it  into  oxaUc  acid  without 
the  production  of  mucic  acid.  The  conditions  of  action 
necessary  to  these  gummy  and  mannitic  ferments  are 
the  same  as  those  which  suit  alcohoHc  ferment.  The 
most  favourable  temperature  is  30°  C.  (86°  F.). 

water  of  flour,  barle3^  and  rice  with  sugar  added.  White  wines  are  more  sub- 
ject to  this  change,  called  ropiness,  than  red  wines  :  as  this  difference  appears 
to  be  connected  with  the  absence  of  tannin  in  the  former,  M.  Francois  pro- 
poses the  addition  of  tannin  to  white  wine,  as  a  remedy  for  the  ropy 
affection. 


CHAPTER  IX. 


LACTIC  FERMENTATION. 

The  lengthened  discussion  into  which  we  have  entered 
with  respect  to  the  history  of  alcohohc  fermentation 
allows  us  to  be  much  more  brief  when  speaking  of  the 
other  phenomena  of  this  order,  which  have  been  gener- 
ally attributed,  since  the  time  of  Pasteur,  to  the  inter- 
vention of  lower  organisms.  Except  the  chemical 
reaction,  and  the  forni  of  the  ferment,  the  general 
characters  of  these  fermentations  are  the  same,  for  the 
very  simple  reason  that  the  conditions  of  nutrition  and 
of  development  of  organic  ferments  do  not  vary  within 
very  wide  limits. 

By  lactic  fermentation  is  understood  the  transforma- 
tion of  certain  sugars,  as  sugar  of  milk  and  grape-sugar 
into  a  sirupy  acid,  soluble  in  water,  (lactic  acid).  By 
comparing  the  formulae  of  the  sugar  which  ferments, 
and  of  the  lactic  acid,  the  only  product  of  its  ferment- 
ation, we  see  that  we  have  here  a  simple  molecular 
transformation,  or  rather  the  splitting  up  of  one  molecule 
into  two  more  simple  equivalent  ones.  We  have,  in 
fact, — 

a  =  2  03 

Sugar.  Lactic  acid. 


1 


194 


ON  FERMENTATION. 


This  transformation  has  been  observed,  long  since, 
'under  many  circumstances. 

M.  Boutron  and  M.  Fremy  were  the  first  to  consider 
it  the  result  of  a  special  fermentation,  independent  of 
the  viscous  fermentation  with  which  it  was  confounded. 
(Ann.  de  Chim.  et  de  Phys.  (3),  vol.  ii.  p.  257,  271.) 

When  milk  turns  sour  spontaneously,  the  sugar  of 
milk  which  it  contains  is  converted  into  lactic  acid,  and 
it  was  from  sour  skimmed  milk  that  Scheele  first 
extracted  lactic  acid,  as  early  as  1780.  Braconnot 
found  the  same  acid  in  rice  left  under  water  in  fermen- 
tation, in  the  juice  of  beetroot,  which,  after  having 
passed  through  the  viscous  fermentation,  and  a  slight 
alcoholic  fermentation,  becomes  sour,  and  produces 
lactic  acid  and  mannite ;  he  also  found  it  in  the  water 
of  fermentation  of  peas  and  boiled  French  beans,  and 
in  the  sour  water  of  baker's  yeast.  The  same  product 
is  found  in  the  sour  water  of  starch  manufacturers,  and 
in  sauerkraut.  M.  Fremy  and  M.  Boutron,  as  well  as 
M.  Pelouze  and  M.  Gelis,  ascertained  the  best  con- 
ditions for  effecting  the  rapid  transformation  of  sugar 
into  lactic  acid.  According  to  their  researches,  lactic 
fermentation  requires  the  presence  of  nitrogenous  albu- 
minoid matter  in  process  of  decomposition,  and  can 
only  continue  if  the  degree  of  acidity  of  the  liquor  is 
kept  from  exceeding  certain  limits.  The  end  is  best 
attained  if  we  saturate  it  from  time  to  time  with  sodium 
carbonate,  or  add  previously  a  quantity  of  chalk,  suffi- 
cient to  neutralize  all  the  acid  which  can  be  formed  at 
the  expense  of  the  sugar. 

We  will  now  give,  in  a  few  words,  the  best  process,  as 
publiiihed  by  the  writers  who  have  studied  this  subject. 


LACTIC  FERMENTATION. 


Three  or  four  litres  of  milk  are  taken,  to  which  from 
200  to  300  grammes  of  sugar  of  milk  is  added,  the 
liquor  is  left  exposed  to  the  air,  in  an  open  vessel,  for 
several  days,  at  a  temperature  between  15°  and  20°  C. 
(50°— 68''  F.).  After  this  time  the  liquor  is  found  to 
be  acid ;  it  is  then  saturated  by  sodium  carbonate,  and 
these  saturations  are  carried  on  successively,  at  inter- 
vals of  twenty-four  hours,  till  all  the  sugar  of  milk  is 
transformed.  It  is  now  boiled,  filtered,  and  cautiously 
evaporated  to  a  sirup.  The  remainder  is  treated  with 
alcohol  at  38°,  which  dissolves  the  sodium  lactate.  This 
solution  is  precipitated  by  a  suitable  quantity  of  sul- 
phuric acid,  which  precipitates  the  soda  ;  it  is  then 
filtered  and  evaporated.  The  remainder,  saturated  with 
lime,  will  give  botryoidal  crystals  of  pure  calcium  lactate. 
(Boutron  and  Fremy.) 

Bensch's  process  is  also  to  be  recommended.  It  consists 
in  dissolving  three  kilogrammes  of  cane-sugar  (about 
6*6  lbs,  av.)  and  15  grammes  (*o83lbs.  av.)  of  tartaric 
acid  in  13  kilogrammes  of  boiling  water.  The  solution 
is  then  left  to  itself  for  some  days ;  after  which  60 
grammes  (•I32lbs.  av.)  of  old  decayed  cheese,  diffused 
in  4  kilogrammes,  (8'81bs.  av.)  of  curdled  and  skimmed 
milk,  i^  kilogrammes  of  washed  chalk  (3*3lbs.  av.)  are 
added.  The  whole  is  put  in  a  warm  place  30° — 35°  C. 
(86'' — 95°  F.),  and  stirred  from  time  to  time.  After 
eight  or  ten  days,  the  liquid  forms  into  a  mass  of  cal- 
cium lactate.  Ten  kilogrammes  of  boiling  water  (22lbs. 
av.)  and  15  grammes  (•033lbs.  av.)  of  slaked  lime  are 
then  added  ;  it  is  boiled,  and  passed  through  a  cloth  ; 
then  evaporated  and  crystallized.  The  crystals  are 
pressed  and  purified  by  new  crystallizations.    The  cal- 


] 


196 


ON  FERMENTATION. 


cium  lactate  is  then  dissolved  in  water  precipitated  by 
a  suitable  quantity  of  sulphuric  acid  (210  grammes  for 
I  kilogramme  of  lactate)  ;  the  calcium  sulphate  is  sepa- 
rated, and  the  liquid  is  neutralized  by  zinc  carbonate  ; 
we  thus  obtain  zinc  lactate,  which  can  be  easily  purified 
by  crystallization,  and  is  readily  converted  by  sulphu- 
retted hydrogen  into  lactic  acid  and  insoluble  zinc  sul- 
phide. Nine  kilogrammes  of  sugar  thus  yield  easily 
io}4  kilogrammes  of  calcium  lactate. 

The  juice  of  fermented  beetroot  (Pelouze  and  J.  Gay- 
Lussac),  and  the  water  in  which  sauerkraut  has  been 
washed  (Liebig),  will  also  serve  for  the  extraction  of 
lactic  acid,  by  analogous  methods,  and  by  the  use  of 
chalk,  or  zinc  carbonate.  The  terms  of  the  reaction, 
and  the  conditions  favourable  to  its  production,  were, 
therefore,  well  known ;  but  before  the  researches  of 
Pasteur,  only  vague  and  ill-founded  ideas  were  held 
respecting  the  cause  which  produced  this  phenomenon. 
The  apparent  absence  of  an  organized  ferment,  which 
had  not  been  found,*  gave  a  powerful  support  to 
Liebig's  theory. 

Casein  or  albuminous  matter  seemed  to  act  upon 
sugar  only  by  its  own  decomposition,  and  it  might  be 
supposed  that  the  molecular  movement,  the  consequence 

*  The  true  lactic  ferment  had,  however,  been  already  surmised  by  Dr.  ke- 
mak,  (Remak  Canstatt's  Jahersbericht,  i.  1841),  and  partly  described  by 
Blondeau  (1848),  "  Journ.  de  Pharm.  "  (3),  vol.  xii,  244  and  336.  Remak  had 
observed  that  the  globules  of  beer-yeast  are  often  mixed  with  smaller  ones, 
which  give  rise  to  a  special  fermentation,  and  can,  in  no  case,  be  transformed 
into  globules  of  the  yeast  of  beer.  Blondeau  points  out  in  beer-yeast  two 
kinds  of  cells,  the  ordinary  ones  of  'oi  millimetre  in  diameter,  setting  up 
alcoholic  fermentation,  and  cells  about  four  times  smaller,  which  give  rise  to 
lactic  fermentation,  liiondeau  attributes  to  them  both  butyric  fermentation 
and  the  ammoniacal  fermentation  of  urea;  but  in  this  he  was  mistaken. 


LACTIC  FERMENTATION.  1 97 

of  this  Spontaneous  decomposition,  was  capable  of 
transmitting  itself  to  the  saccharine  principles  contained 
in  the  liquor. 

M.  Pasteur,  as  early  as  1857,  guided  by  well-founded 
ideas  as  to  the  causes  of  alcoholic  fermentation,  and 
fermentation  in  general,  sought  for  the  lactic  ferment ; 
that  is  to  say,  for  the  organism  which  transforms  sugar 
into  lactic  acid  by  the  effect  of  its  nutrition  and 
development,  and  succeeded  in  finding  and  studying  it, 
although  it  did  not  show  itself  so  distinctly  as  alcoholic 
ferment. 

If  we  examine,"  said  he  (Ann.  de  Chim.  et  de  Phys. 
(3),  vol.  Hi.  p.  407,  Memoire  sur  la  Fermentation  appelee 
Lactique),  with  attention,  ordinary  lactic  fermentation, 
we  find  cases  in  which  we  can  recognize  upon  the 
deposit  of  chalk  and  nitrogeneous  matter,  spots  of  a 
grey  substance  sometimes  forming  a  zone  on  the  surface 
of  the  deposit.  This  substance  is  sometimes  found 
sticking  to  the  upper  parts  of  the  vessel,  whither  it  has 
been  carried  by  the  movement  of  the  gas.  Its  exami- 
nation with  the  microscope  scarcely  enables  us  to 
distinguish  it,  unless  we  are  prepared  for  it  beforehand, 
from  caseum,  disaggregated  gluten,  &c.  ;  so  that  nothing 
shows  that  it  is  a  special  substance,  nor  that  it  has  been 
formed  during  the  fermentation.  Its  apparent  weight 
is  always  very  trifling  compared  with  that  of  the  nitro- 
geneous matter  primarily  necessary  for  the  production 
of  the  phenomenon.  Indeed,  it  is  so  very  often  mixed 
up  with  the  mass  of  caseum  and  chalk,  that  we  should 
have  no  reason  to  suspect  its  existence.  It  is  this, 
nevertheless,  which  plays  the  most  important  part." 
To  prove  this,  M.  Pasteur  thoroughly  washed  fresh 


198 


ON  FERMENTATION. 


yeast  with  fifteen  or  twenty  times  its  weight  of  boiling 
water.  To  this  liquid,  filtered  with  care,  and  containing 
the  most  appropriate  nitrogeneous  and  mineral  aliment 
for  the  nutrition  of  organic  ferments,  was  added  from 
50  to  100  grammes  of  sugar  per  litre,  and  chalk.  A 
trace  of  the  grey  matter  mentioned  above  was  sowed 
in  it,  carbon  dioxide  introduced  instead  of  air,  and  the 
whole  was  kept  at  30°  or  35°  C.  (86^  to  95°  F.).  In  a 
short  time,  all  the  symptoms  of  lactic  fermentation 
w^ere  developed,  accompanied  by  those  of  a  weak  butyric 
fermentation. 

When  the  chalk  has  disappeared,  the  concentrated 
liquid  furnishes  an  abundant  crystallization  of  calcium 
lactate,  while  the  mother  liquor  retains  calcium  buty- 
rate.  Sometimes  the  liquor  becomes  very  viscid.  In 
a  word,  we  have  before  our  eyes  a  most  characteristic 
example  of  lactic  fermentation,  with  all  the  accidents 
and  the  usual  complication  of  this  phenomenon.  At 
the  same  time  that  the  reactions  that  have  been  de- 
scribed take  place,  the  liquid,  limpid  at  first,  becomes 
troubled  ;  a  deposit  appears  which  increases  continually, 
as  fast  as  the  chalk  is  dissolved.    We  may  substitute 


in  this  experiment  any  plastic  nitrogeneous  substance, 
cither  fresh  or  decomposing,  for  the  decoction  of  yeast. 

The  substance  which  is  deposited,  and  whose  produc- 
tion is  correlative  to  the  reactions  known  by  the  name 


Fig.  21. — Lactic  Ferment. 


LACTIC  FERMENTATION. 


199 


of  lactic  fermentation,  exactly  resembles,  when  taken 
in  mass,  ordinary  yeast,  which  has  been  drained  of 
water  and  pressed.  It  is  rather  viscid,  and  of  a  grey 
colour.  Under  the  microscope,  it  appears  as  if  formed" 
of  small  globules  or  short  joints,  either  isolated  or  in 
a  mass,  forming  flakes  resembling  those  of  certain 
amorphous  precipitates  (Fig.  21).  The  globules^  much 
smaller  than  those  of  yeast  of  beer,  are  violently 
agitated  by  the  Brownian  movement,  when  they  are 
isolated.  When  washed  with  much  water  by  decanta- 
tion,  and  then  diffused  in  pure  solution  of  sugar,  it 
begins  to  render  it  acid  at  once,  but  progressively, 
and  very  slowly,  because  acidity  greatly  interferes  with 
its  action.  If  we  add  chalk,  so  as  to  maintain  the 
neutrality  of  the  medium,  the  transformation  of  the 
sugar  is  sensibly  accelerated  ;  at  the  end  of  an  hour, 
gas  begins  to  be  disengaged ;  and  the  liquor  becomes 
charged  with  calcium  lactate  and  butyrate  in  variable 
quantities. 

When  the  solution  of  sugar  contains  nitrogeneous 
matter,  and  salts  suited  to  the  nutrition  of  ferments, 
the  lactic  ferment  is  developed,  and  we  obtain  quanti- 
ties which  have  no  limit  except  the  weight  of  the  sugar 
and  the  albuminous  substances  employed.  But  little 
of  this  ferment  is  necessary  to  transform  a  considerable 
weight  of  sugar.  Its  activity  is  only  weakened  when 
we  dry  it,  or  boil  it  in  water.  This  fermentation  ought 
to  be  set  up  without  access  of  air,  in  order  that  we 
may  not  be  annoyed  by  vegetation  and  infusoria  from 
without. 

Lactic  fermentation,  under  the  conditions  indicated 
by  M.  Pasteur,  %vhe7i  the  lactic  ferment  is  developed 


200 


ON  FERMENTATION. 


sp07ztaneotisly,  is  often  more  rapid  than  alcoholic  fer- 
mentation, and  shorter  than  in  the  ordinary  process. 
(Boutron  and  Fremy,  Bensch,  &c.) 

The  most  favourable  temperature  seems  to  be  about 
35°  C.  (95°  F.). 

If,  without  making  any  change  in  the  conditions  of 
the  saccharine  medium,  which  when  sowed  with  the 
lactic  ferment,  produces  lactic  fermentation,  we  sow 
this  medium  with  globules  of  yeast,  we  observe  the 
appearance  of  alcoholic  fermentation  and  the  develop- 
ment of  alcoholic  ferment. 

Finally,  to  complete  the  analogy  between  the  two 
kinds  of  fermentation,  M.  Pasteur  showed  that  lactic 
ferment  can,  like  alcoholic  ferment,  be  produced  as  if 
spontaneously  in  saccharine  liquids  of  a  composition 
adapted  to  its  development.  Thus  in  alcoholic  fer- 
mentations in  an  artificial  medium,  sowed  with  traces 
of  alcoholic  ferment,  M.  Pasteur  almost  always  saw 
lactic  ferment  and  fermentation  appear  side  by  side 
with  alcoholic  ferment  and  fermentation. 

In  these  cases  of  spontaneous  production  of  lactic,  in 
the  presence  of  alcoholic  ferment,  the  predominance 
of  one  or  the  other  organism,  and  consequently  of  its 
effects,  depends  on  the  composition  of  the  medium, 
being  more  or  less  suited  to  one  or  the  other  ferment, 
and  notably  so,  on  the  neutral  state  of  the  liquid. 

Thus,  if  we  add  magnesia  to  a  mixture  of  solution  of 
sugar  and  beer-yeast,  there  will  be  at  the  same  time 
alcoholic  and  lactic  fermentation  with  precipitation  of 
magnesium  lactate,  and  we  shall  see,  mixed  with  the 
globules  of  yeast  of  beer,  a  considerable  quantity  of 
the   small   globules  of  lactic  ferment.     A  medium, 


LACTIC  FERMENTATION. 


201 


slightly  alkaline,  is  more  suited  to  the  development 
of  the  new  ferment ;  it  is  the  same  with  respect  to 
alcoholic  ferment. 

The  concomitant  development  of  alcoholic  and  lactic 
ferment  explains  why  frequently  the  presence  of  lactic 
acid  has  often  been  observed  as  one  of  the  products  of 
the  decomposition  of  sugar  under  the  influence  of  becr- 
yeast. 

Most  careful  and  frequent  experiments  proved  to 
M.  Pasteur,  that  not  even  the  smallest  quantity  of 
lactic  acid  is  formed  in  normal  alcoholic  fermentation. 

When  it  does  appear,  which  is  very  rarely,  unless  we 
choose  favourable  conditions,  we  may  be  sure  that  the 
beer-yeast  is  mixed  with  lactic  ferment,  which  can 
easily  be  recognized  by  its  form,  its  size,  and  its 
movement. 

To  be  certain  of  the  presence  or  absence  of  lactic 
acid,  we  operate  as  has  been  before  explained  (page  30) 
when  treating  of  succinic  acid  and  glycerine.  The 
ethereal  alcoholic  solution  of  the  extract  of  the  fer- 
mented liquid  is  evaporated  ;  the  remainder  is  saturated 
with  lime-water ;  the  liquid  is  again  evaporated,  and 
the  remainder  is  taken  up  by  ethereal  alcohol.  The 
residuum  is  finally  heated  with  boiling  alcohol  at  95  per 
cent,  which  dissolves  the  calcium  lactate. 

The  following  substances  are  susceptible  of  undergoing 
lactic  fermentation,  the  glucoses  and  those  bodies  which 
are  convertible  into  glucoses.  The  saccharoses  which 
most  easily  undergo  alcoholic  fermentation,  are  those 
which  are  with  the  greatest  difficulty  transformed  into 
lactic  acid,  and  vice  versa.  Sugar  of  milk  does  not  readily 
produce  alcohol,  while  it  is  much  more  easily  resolved 


202 


ON  FERMENTATION. 


into  lactic  acid,  although  according  to  Lubolt  (Journ. 
J.  Prakt.  Chim.  vol.  77,  p.  282)  and  Proust  (Ann.  de 
Chim.  et  Phys.  (2),  vol.  10,  29)  the  action  is  very  slow, 
sugar  of  milk  being  found  in  the  liquor  up  to  the  end 
of  the  fermentation. 

Sorbin,  inosite,  mannite,  and  dulcite  do  not  un- 
dergo alcoholic  fermentation,  but  are  affected  by  lactic 
ferment. 

Calcium  malate,  and,  in  general,  all  substances  whose 
fermentation  produces  butyric  acid,  seem  to  have  the 
power  of  yielding  lactic  acid. 

In  the  fermentation  of  calcium  malate,  carbon  dioxide 
is  disengaged. 

C4  H6  o^   =        H«  0=^    +    C  02. 
Malic  Acid.  Lactic  Acid.    Carbon  dioxide. 


CHAPTER  X. 


AMMONIACAL  FERMENTATION. 

Urea  (C  O)  is,  as  is  well  known,  one  of  the 

most  important  constituents  of  urine.  It  appears  in  it 
as  the  principal  form  under  which  nitrogen  is  eliminated 
from  the  animal  organism. 

This  body,  of  simple  composition,  crystallizable  in 
long  prisms,  only  differs  from  ammonium  carbonate  by 
the  elements  of  water. 

We  have,  in  fact, — 

(CH^  N2  O)   4-   H^O     =     C02     +     2   N  H\* 
Urea.  Water.       Carbon  dioxide.  Ammonia. 

By  boiling  in  alkaline,  or  even  in  pure  water,  this 
phenomenon  of  hydratation  is  produced  more  or  less 
quickly.  We  even  observe  it  with  the  more  complex 
compounds,  known  under  the  name  of  ureids,  and  in 
which  we  must  admit  that  the  molecule  of  urea  is 
associated  with  other  organic  groups,  such  as  uric  acid, 
alloxane,  creatine,  &c. 

A  solution  of  pure  urea  in  water  may  be  preserved 
for  a  long  time  without  change.  It  is  not  so  with  the 
natural  solutions  of  urea,  such  as  urine,  which  contains 

*  Or  better,  CH^      O  +  2      O  =  (NH*)^  C  O^. 

Urea.  Water.      Ammonium  carbona. 


204 


ON  FERMENTATION. 


salts  and  other  nitrogeneous  principles  more  similar  to 
albuminous  substances. 

It  is  well  known  that,  after  a  longer  or  shorter  time, 
according  to  the  conditions  of  temperature,  and  the 
state  of  health  of  the  individual  who  has  excreted 
the  urine,  this  liquid,  after  its  emission,  becomes  alka- 
line, instead  of  acid  as  it  was  before ;  at  the  same  time, 
it  exhales  a  very  decided  odour  of  ammonia ;  at  this 
moment  the  urea  has  disappeared,  or  is  on  the  point  of 
disappearing  entirely;  we  find  in  its  place  an  equivalent 
quality  of  ammonium  carbonate. 

In  certain  cases,  the  urine  is  already  alkaline  and 
ammoniacal,  when  in  the  bladder. 

The  name  of  ammoniacal  fermentation  has  been 
given  to  this  spontaneous  transformation.  (Dumas, 
Traite  de  Chimie,  vol.  6,  p.  380.) 

According  to  the  observations  of  Muller  (i860,  Journ. 
fiir  Prakt.  Chem.,  81,  p.  467)  and  of  Pasteur  (Comp. 
Rend.,  vol.  50,  p.  869,  May,  i860),  the  transformation  of 
urea  into  ammonia  and  carbon  dioxide  is  due  to  the 
intervention  of  a  special  organic  ferment,  produced  by 
one  of  the  toriilacei  and  formed  of  chaplets  of  globules 
very  similar  to  those  of  beer-yeast,  but  much  smaller ; 
their  diameter  is  about  5-0V0  of  a  millimetre  (,0000078  in.). 
M.  Van  Tieghem  has  very  thoroughly  studied  this  fer- 
ment. It  is  found  in  the  white  deposit  left  at  the  bottom 
of  urinals.  M.  Jaquemart  (Ann.  Chim.  Phys.  (3),  vol.  7, 
p.  149,  1843)  had  already  noticed  that  this  deposit  was 
very  apt  to  excite  this  transformation. 

Long  study  of  the  organic  products  which  are  de- 
veloped in  urine  exposed  to  air,  has  convinced  M.  Van 
Tieghem  of  the  constant  presence  of  a  (torulaccous) 


AMMONIACxVL  FERMENTATION. 


205 


ferment,  whenever  urea  ferments ;  and  of  the  mtimate 
connection  which  exists  between  its  easy  or  difficult 
development,  and  the  rapid  or  slow  transformation  of 
the  urea. 

"  In  the  case,  seldom  realized,  in  which  this  torulaceous 
growth  is  developed  alone,  the  liquid  remains  lim.pid, 
the  fermentation  is  rapid,  and  the  deposit  which  forms 
at  the  bottom  of  the  vessel  is  composed  exclusively  of 
chaplets  and  masses  of  globules  mixed  with  crystals  of 
urates  and  of  ammonium-magnesium  phosphate.  If  the 
torulaceous  growth  is  only  accompanied  by  infusoria, 
as  is  usually  the  case,  the  fermentation,  though  some- 
what slower,  is  still  easy;  but  if  there  appear,  besides 
the  infusoria,  vegetable  productions  in  the  liquid  and  on 
the  surface,  the  torulaceous  growth  is  developed  with 
difficulty,  and  the  transformation  is  very  slow ;  the 
liquid  may  remain  acid  or  neutral  for  months  together. 

"  If,  instead  of  leaving  the  urine  to  the  variable  chances 
which  the  sequence  of  the  presence  of  the  germs  in  the 
air  involve,  we  place  it  in  a  stove  in  a  corked  bottle, 
having  added  to  it  a  trace  of  the  deposit  of  a  good  fer- 
mentation, all  accidental  variations  disappear,  and  the 
phexiomenon  takes  place  always  in  the  same  manner ; 
one  or  two  days  are  sufficient  for  the  urea  to  disappear, 
and  at  the  same  time  the  torulaceous  growth  alone  is 
developed. 

"  The  transformation  of  urea  in  urine  is  therefore 
correlative  to  the  life  and  development  of  an  organic 
vegetable  ferment.  This  ferment  is  developed  within 
the  liquid  itself,  and  especially  at  the  bottom  of  the 
vessel,  where,  by  its  accumulation,  it  forms  a  whitish 
deposit,  and  is  composed  of  chaplets,  or  small  masses  of 


206 


ON  FERMENTATION. 


spherical  globules,  without  granulations,  without  any 
distinct  envelope  of  the  contents,  and  which  appear  to 
develop  by  budding;  their  diameter  is  about  '0015 
millimetres  ('000059  in.)." 

M.  Van  Teighem  considers  that  he  has  proved  by 
direct  experiment,  that  the  splitting  up  of  hippuric  acid 
by  hydratation,  into  benzoic  acid  and  glycocol,  which  is 
observed  in  the  urine  of  the  herbivora  after  emission,  is 
due  to  a  fermentation  analogous  to  that  which  splits  up 
the  urea.  The  active  ferment  must  be  identical  with 
the  ammoniacal  ferment.  Thus,  ammonium  hippurate 
dissolved,  either  in  yeast-water  or  in  solution  of  sugar 
containing  phosphates,  is  always  split  up  in  consequence 
of  the  development  of  a  microscopic  vegetable  organ- 
ism identical  with  the  torulaceous  growth  described 
above.  (Comp.  Rend,  de  TAcad.  des  Sci.,  vol.  58,  p.  533.) 

According  to  Miiller,  the  activity  of  the  phenomenon 
is  proportionate  to  the  number  of  globules.  When  to 
a  mixture  of  sugar  and  urea,  dissolved  in  water,  we  add 
beer-yeast,  we  always  see  the  small  globules  of  ammo- 
niacal ferment  make  their  appearance,  as  soon  as  the 
liquid  shows  an  alkaline  reaction  (Pasteur).  The  yeast 
of  beer,  by  itself,  decomposes  the  sugar  without  exciting 
the  decomposition  of  the  urea.  The  most  suitable 
temperature  is  that  of  the  human  body,  37°  C.  (99  F''). 

Finally,  as  is  the  case  with  the  must  of  grapes  and 
the  wort  of  beer,  the  ferment  does  not  pre-exist  in  the 
urine  ;  it  must  be  brought  from  without,  in  the  form 
of  germs. 

The  necessity  for  ammoniacal  ferment,  in  order  to 
transform  the  urea  found  in  urine,  under  ordinary  con- 
ditions of  temperature,  raises  a  medical  question  of 


AMMONIACAL  FERMENTATION.  20/ 

great  importance.  In  fact,  many  cases  of  disease  have 
been  observed  in  which  the  urine  was  more  or  less  am- 
moniacal  in  the  bladder.  This  change  usually  accom- 
panies either  vesical  or  renal  injuries,  or  serious  general 
diseases,  such  as  typhoid  fever.  When  the  urine  is  am- 
moniacal  after  the  catheter  has  been  introduced,  it  is  to 
be  feared  that  the  instrument  has  served  as  a  vehicle, 
and  carried  into  the  bladder  the  germs  of  the  torula- 
ceous  growth,  thus  setting  up  infection.  Ammoniacal 
urine,  from  a  patient  at  la  Charite,  was  examined  by 
M.  Gayon,  at  the  very  instant  of  emission  ;  he  found  in 
it  innumerable  organisms  :  this  examination  took  place 
some  days  after  the  patient  had  been  relieved  by  the 
catheter.  However,  in  the  opinion  of  surgeons,  the 
urine  often  shows  itself  to  be  ammoniacal  without  any 
previous  operation  of  this  kind  ;  for  example,  in  cases 
of  retention,  although  the  long  continuance  of  the 
liquid  in  the  bladder  may  not  be  the  only  condition  of 
this  phenomenon.  In  fact,  M.  Verneuil  did  not  find 
the  urine  ammoniacal  in  the  case  of  an  hysterical  young 
girl,  in  whose  case  the  catheter  had  been  passed,  after 
several  days*  retention. 

As  M.  Pasteur  has  observed,  the  canal  of  the  urethra, 
with  relation  to  the  infinitely  small  germs  which  act  as 
ferments,  may  be  compared  to  the  Thames  tunnel, 
through  which  they  can  pass  and  repass  easily.  If  then, 
we  should  feel  astonished  at  anything,  it  is  rather  at 
finding  the  ammoniacal  infection  occur  so  infrequently. 
It  is,  perhaps,  because  the  urine  is  usually  acid,  and 
that  this  acid  is  unfavourable  to  the  development  of  the 
germs  of  fermentation. 

However  this  mav  be,  M.  Pasteur,  without  being 


208 


ON  FERMENTATION. 


able  to  give  any  positive  proof,  is  inclined  to  believe 
that  the  decomposition  of  urea  is  not  possible,  without 
the  presence  of  a  peculiar  ferment,  and  that  there  is 
no  purely  chemical  reaction  in  the  human  body,  capable 
of  giving  rise  to  ammonium  carbonate  in  the  urine. 
This  conviction  has  been  the  result  of  long  study  of 
ferments,  and  of  his  great  experience.  He  owns, 
how^ever,  that  the  definite  solution  has  not  yet  been 
obtained.  {See  on  this  subject  the  discussion  on  am- 
moniacal  urine  at  the  Academic  de  Medicine,  Moniteur 
Scientifique  (3),  vol.  9,  p.  143.) 

In  conclusion,  the  transformation  of  the  urea  in  urine, 
at  the  ordinary  temperature,  may  certainly  be  caused 
by  a  special  ferment.  It  is  possible,  but  as  yet  not  cer- 
tainly proved,  that  the  presence  of  this  ferment  may  be 
indispensable. 


CHAPTER  XI. 


BUTYRIC  FERMENTATION  AND  PUTREFACTION. 

A  GREAT  number  of  chemical  compounds  are  sus- 
ceptible of  fermenting  butyrically,  that  is  to  say,  yield- 
ing butyric  acid  as  a  product  of  their  transformation, 
when  they  are  placed  under  suitable  conditions. 

Such  are  lactic  acid,  and  all  substances  capable  of 
undergoing  lactic  fermentation — sugars,  amylaceous 
matter,  tartaric,  citric,  malic,  mucic  acids,  and  al- 
buminoid substances. 

It  is  easy  to  represent  by  equations  the  production 
of  butyric  acid  at  the  expense  of  the  greater  part  of 
these  bodies,  which  are  well  defined  and  of  well-known 
composition. 

Thus  sugar  and  lactic  acid  would  give — 

a     o^  =  2  a  H6  03  ==  a     o'^  +    2  co^  + 

Glucose.         Lactic  acid.       Butyric  acid.       Carb.  dioxide.  Hydrogen. 

For  malic  acid  we  shall  have — 

2  O^  =  2  a       03  +  2  CO2    =   G*      O*  + 

Malic  acid.  Lactic  acid.     Carbon  dioxide.    Butyric  acid. 

4  CO2  + 
Carb.  diox.          Hydrogen.  , 

Tartaric  acid  would  be  resolved   into   butyric  acid, 
carbon  dioxide,  and  water : 
10 


210 


ON  FERMENTATION. 


2  C*        06  =  a      02    +      4  C02       +  2  O 
Tartaric  acid.       Butyric  acid.       Carbon  dioxide.  Water. 

Perhaps  lactic  acid  is  formed  first : 

3  0«  =2      H«  O^  +    6  C02  -|.  H« 

Tartaric  acid.  Lactic  acid.       Carb.  diox.  Hydrogen. 

According  to  M.  Personne,  citric  acid  would  be  trans- 
formed previously  into  lactic  and  acetic  acid  : 

4  O?  +  2  H2  O  ==  3  02  +  4  C3      03  +  6  CO  2 

Citric  acid.  Water.         Acetic  acid.  Lactic  acid,      Carb.  diox. 

Mucic  acid,  which  differs  from  citric  acid  only  by 
containing  more  water,  is  resolved,  like  it,  into  acetic 
and  butyric  acids,  carbon  dioxide,  and  hydrogen : 

3  C«       O^  =  3  C2      02  +  C^      02  +   8  CO2  +  U^^ 

Mucic  acid.  Acetic  acid.       Butyric  acid.      Carb.  diox  Hydrogen. 

In  other  analogous  phenomena,  which  go  on  at  the 
expense  of  the  same  substances,  we  find  acids  homo- 
logous to  butyric  acid  ;  in  the  same  way  as,  in  alcoholic 
fermentation,  alcohols  are  formed  with  higher  equiva- 
lents than  that  of  ethyl-alcohol. 

Thus,  under  the  conditions  of  lactic  fermentation,  we 
see  the  formation  of  propionic  acid,  acetic  acid,  and 
valerianic  acid  at  the  expense  of  sugar  or  of  starch. 

Glycerine,  placed  in  contact  for  a  long  time  with 
beer-yeast,  also  furnishes  propionic  acid,  mixed  with 
formic  and  acetic  acids.  M.  Monoyer  represents  in 
a  general  manner  the  production  of  fatty  acids  at  the 
expense  of  sugar  by  the  equation — ■ 

(C«  II'2  6« )  =  [C"  H-n  02J  +  (n  —  2)  CH*^  O^ 

3 

Fatty  acid.  Formic  acid. 


J 
i 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  211 


Formic  acid  itself  would  be  decomposed  into  carbon 
dioxide  and  hydrogen. 

Crude  calcium  tartrate,  mixed  with  organic  matter, 
and  left  under  water,  in  summer,  ferments  and  produces 
an  acid  which  was  at  first  identified  with  propionic  acid, 
but  which,  according  to  Limprecht  and  Von  Uslar,  must 
be  isomerous  with  ordinary  propionic  acid. 

Crude  tartar,  without  the  addition  of  lime,  gives 
nothing  but  acetic  acid.    We  have — 

2  C^       06  =  C3  H«  O^  +  5  CO^  + 
Tartaric  acid.      Propionic  acid.  Carb.  diox.  Hydrogen. 

C^  H«  O^  =  C^       O^  +  2  CO^  +  H*^ 

Acetic  acid. 

In  the  fermentation  of  mucic  acid,  of  which  we  have 
spoken  above,  the  butyric  acid,  which  only -appears 
slowly,  and  in  small  quantity,  ought  to  be  considered  as 
a  secondary  product.  The  principal  phenomenon  may 
be  summed  up  in  a  chemical  point  of  view  by  the 
equation — 

C6        O^  =  2  C2       02  4-  2  C02  +  H2 

All  the  compounds  of  the  malic  group,  such  as  malic, 
fumaric,  aconitic,  aspartic  acids,  and  asparagine,  undergo, 
under  the  form  of  salts  of  lime,  and  in  the  presence 
of  animal  matter  in  process  of  decomposition,  a  trans- 
formation into  different  products,  among  which  we  may 
consider  succinic  acid  as  the  principal  term. 
With  malic  acid,  we  should  have — 

2  C^  H6  05  =       H6  O^  +  C2       02  +  2  C02  + 
Malic  acid.         Succinic  acid.   Acetic  acid.     Carb.  diox.  •  Hydrogen. 

The  sum  of  the  last  three  terms  of  the  second  member 


212 


ON  FERMENTATION. 


gives  the  composition  of  tartaric  acid  ;  we  may,  there- 
fore, consider  succinic  fermentation  as  the  result  of  two 
reactions.  In  the  first,  two  molecules  of  malic  acid  are 
converted  into  one  molecule  of  tartaric  acid,  and  one  of 
succinic  acid : 

2  O       O^  =  a       O^  +  O  O^ 

Malic  acid.        Succinic  acid.    Tartaric  acid. 

The  second  reaction  would  be  the  fermentation  of  tar- 
taric acid  formulated  above. 

The  production  of  valerianic  acid  in  the  succinic 
fermentation  of  calcium  malate  is  explained  by  the 
equation — 

2  (C*       O^)  =  0^  +  3  C02  +  H2 

Succinic  acid.    Valerianic  acid. 

As  maleic,  fumaric,  and  aconitic  acids  only  differ  from 
malic  acid  by  less  water,  their  transformation  into 
succinic  acid  is  explained  in  the  same  manner. 

As  aspartic  acid  may  be  considered  as  amido-suc- 
cinic  acid,  and  asparagine  as  amido-succinamic  acid, 
the  production  of  succinic  acid  at  their  expense  is  not 
surprising. 

We  have  just  passed  in  review  a  great  number  of 
reactions  ;  they  all  belong  to  the  class  of  fermentation, 
because  they  are  excited  by  certain  bodies  which  act 
only  by  their  presence. 

They  have,  besides,  one  thing  in  common — the  forma- 
tion of  acids  of  the  fatty  series. 

We  must  consider  it  as  yet  vague  and  uncertain 
whether  the  cause  of  these  fermentations  is  to  be  attri- 
buted to  the  presence  of  a  special  ferment  for  each,  as 
\vc  have  seen  in  lactic  and  alcoholic  fermentations. 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  21 3 


In  fact,  this  cause  has  only  been  thoroughly  studied 
and  determined  by  Pasteur,  for  the  butyric  fermentation 
of  sugar  and  calcium  lactate. 

In  the  fermentation  of  ammonium  tartrate,  M.  Pas- 
teur found  a  ferment  similar  to  lactic  ferment,  which, 
acting  on  ammonium  paratartrate,  causes  the  decom- 
position of  the  levo-tartrate  ;  which  affords  a  new 
example  of  elective  fermentation,  comparable  to  that 
which  we  have  observed  in  levulose  and  glucose. 

In  M.  Pasteur's  opinion,  the  butyric  decomposition 
of  calcium  lactate  is  due  to  the  presence  in  the  liquid 
of  a  special  itrmQwt—fermeiitum  butyrictcm. 

"  Butyric  ferment  is  composed  of  little  cylindrical 
rods,  rounded  at  the  extremities,  usually  straight,  either 
isolated  or  united  in  a  chain  of  two,  three,  or  four  joints, 
and  even  of  more.  The  diameter  of  these  small  rods 
is  generally  i-^^o  of  a  millimetre,  and  the  length  of  the 
isolated  portions  from  yo%o  tu^o  i^m-  (•0000687  to 
'000687  in.).  These  organisms  move  forward  by  sliding. 
During  this  movement  their  body  remains  rigid  or 
undulates  slightly  ;  they  spin  round,  they  balance  them- 
selves on  end,  and  agitate  their  extremities:  they  are  often 
bent.  These  singular  organisms  are  reproduced  by  fission. 

"  The  butyric  ferment  is,  therefore,  an  infusorium  of 
the  genus  vibrio."  (Pasteur,  Comp.  Rend.,  52,  p.  344, 
February,  1861). 

The  same  writer  has  ascertained  that  this  ferment, 
placed  in  a  solution  of  sugar  containing  phosphates  and 
ammoniacal  salts,  reproduces,  and  causes  butyric  fer- 
mentation. 

The  conditions  of  its  development  are  similar  to  those 
of  lactic  fermentation. 


I 

i 


214 


ON  FERMENTATION. 


The  most  favourable  temperature  is  40°  C.  (104''  F.) : 
the  medium  should  be  neutral^  or  slightly  alkaline.  An 
acid  medium  is  opposed  to  the  development  of  the 
germs  of  butyric  fermentation.  However^  when  once 
formed,  they  can  live  and  excite  the  decomposition 
of  sugar  or  lactic  acid  in  an  acid  m.edium,  provided 
there  be  no  excess  of  acidity. 

M.  Pasteur  at  first  asserted  that  butyric  vibrios 
not  only  lived  without  free  oxygen,*  but  that  oxygen 
kills  them.  The  respiratory  theory  of  fermentation, 
proposed  by  this  observer,  does  not  agree  with  this  fact. 
If  fermentation  is  the  result  of  such  a  need  of  oxygen, 
that  the  ferment  takes  it  up  from  organic  compounds, 
exciting  their  decomposition  by  a  rupture  of  equilibrium, 
we  cannot  understand  how  oxygen  can  act  as  a  poison 
to  the  ferment.  I  do  not  know  whether  M.  Pasteur 
has  since  maintained  the  opinion  that  oxygen  kills 
butyric  vibrios. 

The  conditions  of  nutrition  of  butyric  ferment  are, 
according  to  Pasteur,  the  same  as  those  of  ferments  in 
general.  However,  considering  its  tardy  appearance, 
as  compared  with  lactic  ferment,  in  mixtures  which 
undergo  lacto-butyric  decomposition,  we  may  admit 
that  it  requires  albuminoid  substances  in  a  process  of 
more  advanced  change  for  its  nourishment. 

In  sugar,  we  generally  see  viscous,  lactic,  and  butyric 
fermentation  appear  in  succession.  However,  it  is  not 
proved  that  butyric  fermentation  cannot  appear  before 
lactic  fermentation,  and  thus  act  on  the  sugar  itself. 

Putrefaction — Putrid  Fermentation, — Albuminoid  sub- 
stances, and  bodies  allied  to  them,  which  enter  into  the 

*  Tlic  alcoholic  and  lactic  ferments  arc  in  the  same  category. 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  21$ 

composition  of  living  organisms,  have,  for  a  long  time, 
enjoyed  a  special  reputation  for  instability,  which  varies, 
however,  according  to  the  nature  of  the  substance. 
Before  Pasteur's  researches,  it  was  generally  admitted, 
that  as  soon  as  the  influence  of  life  is  withdrawn  (the 
vital  force  which  alone  was  able  to  maintain  them  in 
their  integrity),  these  products  begin  to  be  transformed, 
to  change,  and  to  be  decomposed  into  several  principles, 
among  which  are  found  compounds  with  a  strong  and 
putrid  odour. 

The  remarkable  researches  of  Appert  on  the  preserva- 
tion of  animal  substances,  and  those  of  Gay-Lussac  on 
the  fermentation  of  the  must  of  grapes,  had  given  the 
idea  that  the  momentary  intervention  of  oxygen  is 
necessary  to  excite  the  first  step  in  this  decomposition. 
This  initial  impulse  once  given,  the  phenomenon  of 
decomposition  goes  on  spontaneously ;  and  the  organic 
matter  in  process  of  transformation  is  even  susceptible 
of  transmitting  the  molecular  movement  with  which  it 
is  imbued  to  more  stable  bodies,  such  as  sugar,  which 
of  themselves  undergo  no  modification.  This  is,  as  we 
have  already  seen,  the  theory  of  fermentation  borrowed 
by  Liebig  from  Stahl  and  Willis,  with  certain  modifica- 
tions of  form. 

However,  Schwann  (Ann.  de  Poggend.,  41,  p.  184), 
Ure  (Journ.  Prakt.  Chem.,  19,  186),  and  Helmholtz  (J.  F. 
Prakt.  Chem.,  31,  p.  429),  had  shown  that  the  greater  part 
of  bodies  subject  to  decomposition,  when  heated  in  a 
retort  with  water,  so  as  to  drive  out  all  the  air  by  boiling, 
are  no  longer  decomposed,  if  instead  of  allowing  ordi- 
nary air  to  enter  the  retort  as  it  grows  cold,  we  are 
careful  only  to  admit  air  previously  subjected  to  a  red 


2l6 


ON  FERMENTATION. 


heat.  Under  these  conditions,  putrefaction  does  not 
make  its  appearance,  and  we  no  longer  observe  the 
development  of  infusoria  and  mildews. 

The  abundant  presence  of  infusoria  and  mildews  in 
putrefaction  had  been  long  known,  but  it  was  not  thought 
that  these  microscopic  beings  were  the  true  causes 
which  determined  the  decomposition.  They  are  de- 
veloped, it  was  said,  owing  to  germs  brought  by  the 
air,  or  already  contained  in  the  decomposing  bodies,  or 
by  spontaneous  generation,  and  because  they  find  a  soil 
favourable  to  their  nutrition.  The  bond  between  their 
appearance  and  putrefaction  was  only  one  of  con- 
comitance. 

Schwann,  and  the  other  authors  quoted  above,  thought, 
on  the  contrary,  that  the  germs  of  infusoria  and  of  mil- 
dews set  up  putrefaction  by  their  development ;  and,  as 
a  proof  of  this,  they  brought  forward  their  experiments, 
in  which  putrefaction  was  no  longer  produced,  when  the 
pre-existent  germs  were  destroyed,  and  their  introduction 
by  means  of  the  air  was  prevented. 

As  the  calcination  of  the  air  gave  rise  to  some  objec- 
tions, especially  to  that  of  a  possible  change  in  the 
constituent  principles  of  this  gaseous  mixture,  Schroeder 
and  Th.  V.  Dusch  (Ann.  der  Chem.  und  Pharm.,  vol.  89, 
p.  232),  repeated  Schwann's  experiments,  with  this 
difference,  that  instead  of  allowing  calcined  air  to  re- 
enter the  retort  in  which  the  organic  substance  had  been 
boiled,  they  simply  filtered  the  air  through  a  sufficiently 
thick  Tayer  of  cotton  wool  ;  they  thus  succeeded  in 
mechanically  arresting  the  germs  and  solid  matters  held 
in  suspension,  but  without  influencing  in  any  way  the 
properties  and  the  composition  of  the  air.    The  wort  of 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  21/ 

beer,  broth,  meats  recently  boiled  in  water,  are  then 
preserved  very  well,  even  during  the  heat  of  summer. 

Some  contradictory  facts,  however,  lent  support  to 
the  arguments  of  the  adversaries  of  the  theory  of  putre- 
faction under  the  influence  of  infusoria.  Thus,  the 
authors  of  the  before-mentioned  experiments  had  them- 
selves ascertained  that  milk  recently  boiled  coagulates, 
grows  sour,  and  putrefies,  just  as  well  in  air  that  has 
been  strained  as  in  ordinary  air  ;  meat  not  steeped  in 
water,  but  simply  heated  in  a  water-bath,  is  also  not 
preserved  in  strained  air :  in  these  two  cases  we  observe 
neither  infusoria  nor  mildew,  and  yet  decomposition 
is  produced. 

It  is  then  evident,  it  was  said  (Gerhardt,  Chimie 
Organ.,  vol.  4,  pp.  545),  that  it  is  indeed  the  air  which 
brings  and  deposits  in  matters  in  a  state  of  putrefac- 
tion the  germs  of  organisms,  but  it  is  not  less  certain 
that  these  are  not  the  first  cause  of  decomposition,  since 
it  can  be  produced  without  their  intervention.  If  the 
calcined  or  strained  air  is  less  active,  in  many  experi- 
ments, than  ordinary  air,  it  is  because  not  only  are  the 
germs  of  infusoria  removed  by  these  operations,  but 
also  the  remains  of  decomposed  matter  which  are  sus- 
pended in  it ;  that  is  to  say,  ferments  whose  activity 
would  be  added  to  that  of  oxygen. 

The  question  was  in  this  state  when  Pasteur  resumed 
the  study  of  putrefaction,  by  looking  upon  it  in  the  same 
light  as  had  guided  him  in  his  researches  on  fermenta- 
tion. Sustained  by  the  idea  that  all  these  phenomena 
can  be  explained  by  the  presence,  the  development, 
and  the  multiplication  of  microscopical  plants  or 
animals,  he  sought  to  prove  that  some  of  these  exist 


2l8 


ON  FERMENTATION. 


also  in  the  putrefaction  of  animal  nitrogenous  sub- 
stances. 

These  experiments  were  conducted  by  two  methods 
which  lead  to  the  same  end,  and  confirm  each  other. 

On  the  one  hand,  they  tend  to  show  that  putrefaction 
is  always  accompanied  by  the  presence,  the  develop- 
ment, and  the  multiplication  of  infinitely  small,  organ- 
ized, living  beings  ;  on  the  other  hand,  they  prove, 
that  whenever  we  place  ourselves  under,  conditions 
calculated  to  avoid  the  presence  of  the  germs  of  organ- 
isms, at  the  commencement  of  the  experiment,  decom- 
position does  not  take  place,  even  in  prodtLcts  the  most 
liable  to  it. 

By  their  precision  and  their  extent,  they  are  calcu- 
lated to  remove  the  objections  raised  by  the  partial 
decompositions  noticed  by  his  predecessors,  Schroeder 
and  V.  Dusch,  and  which  we  have  mentioned  before. 

We  will  speak  of  the  second  series  of  experiments 
when  we  treat  of  the  origin  of  ferments,  only  saying 
here,  that  the  many  trials  made  by  Pasteur  lead  to  a 
positive  solution  of  the  question.  By  preventing  con- 
tact of  the  germs  with  animal  matter,  we  prevent,  at 
the  same  time,  every  trace  of  fermentation  from  showing 
itself. 

M.  Pasteur  distinguishes  two  orders  of  phenomena 
in  putrefaction  ;  some  are  produced  under  the  influence 
of  organic  ferments  which  live  without  the  aid  of  oxygen, 
like  butyric  ferment ;  in  others,  on  the  contrary,  the 
oxygen  takes  part,  as  an  essential  element,  promoting 
combustion  ;  oxidation  is  also  excited  by  organisms. 

We  shall  treat  of  slow  combustions  in  the  chapter 
on  acetic  fermentation  ;  this  will  give  us  a  simple  and 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  2ig 

clear  example  of  the  reactions  of  this  group,  and  may 
be  considered  as  the  type  of  slow  combustions.  We 
have  nothing  to  add  here  to  what  will  be  said  on  this 
subject,  and  there  only  remains  for  us  to  speak  of  putre- 
factions without  oxygen,  or  putrid  fermentation. 

When,  in  a  putrescible  liquid,  containing  albuminoid 
organic  matter,  the  dissolved  oxygen  has  been  absorbed, 
and  has  completely  disappeared  under  the  influence  of 
the  first  infusoria  developed,  such  as  the  Monas  crepns- 
cnlum,  and  the  Bacterium  termo,  the  vibrio  ferments, 
which  do  not  require  this  gas  to  sustain  their  life,  begin 
to  show  themselves,  and  putrefaction  is  immediately  set 
up.  It  is  accelerated  by  degrees,  following  the  progres- 
sive increase  of  the  vibrios.  As  to  the  putridity,  it 
becomes  so  intense,  that  the  examination  of  a  single 
drop  of  the  liquid,  under  the  microscope,  is  a  very  painful 
task." 

"  It  follows,  from  what  has  been  said,  that  contact  of  air 
is  by  no  means  necessary  for  the  development  of  putre- 
faction. On  the  contrary,  if  the  oxygen  dissolved  in  a 
putrescible  liquid  was  not  at  once  removed  by  the  action 
of  special  organisms,  putrefaction  would  not  take  place  ; 
the  oxygen  would  destroy  the  vibrios  which  would  try 
to  develop  at  first." 

When  the  putrescible  liquid  is  exposed  to  the  air,  we 
notice  the  two  kinds  of  reactions  simultaneously ;  there 
forms  on  the  surface  a  complete  film,  composed  of 
bacteria,  mucors,  and  mucidines,  which  excludes  the 
oxygen,  and  prevents  its  penetrating  into  the  liquid. 
The  vibrios  which  multiply  there,  under  shelter  of  this 
rampart,  transform  by  fermentation  the  albuminoid 
matter  into  more  simple  products,  while  the  bacteria 


220 


ON  FERMENTATION. 


and  mucors  excite  the  combustion  of  these  products, 
and  bring  them  back  to  the  state  of  the  least  complex 
chemical  combinations.  Such  is  the  representation  of 
the  whole  of  the  phenomena  of  putrefaction  as  drawn 
by  M.  Pasteur  (Comp.  Rend.,  June,  1863). 

We  have  already  made  some  reservations  relative  to 
this  singular  property  of  vibrios — that  of  not  being  able 
to  endure  the  presence  of  oxygen. 

The  opinion  held  by  Schwann,  Ure,  Helmholtz, 
Schroeder,  and  V.  Dusch,  and  finally  by  Pasteur,  rela- 
tive to  the  cause  of  putrefaction,  is  corrobor^lted  by  the 
very  process  which  is  employed  to  preserve  perishable 
bodies.  The  conditions  of  preservation  are  precisely 
such  as  oppose  the  development  of  organisms. 

Such  are  the  employment  of  cold,  zero  C.  (32°  F.),  and 
below  ;  also  of  a  sufficiently  high  temperature.  Cooked 
albuminous  matter  resists  putrefaction  much  longer, 
because  the  germs  which  were  there  are  destroyed  ;  but 
decomposition  will,  nevertheless,  show  itself  if  we  do 
not  carefully  guard  against  effects  from  without.  Ap- 
pert's  process,  which  consists  in  cooking  meat,  or  other 
perishable  substances,  in  iron  boxes  hermetically  sealed, 
realizes  these  conditions.  The  germs  are  killed,  and 
there  is  no  possibility  of  fresh  ones  entering.  As,  at  the 
same  time,  the  small  quantity  of  air  contained  in  the 
box  loses  its  oxygen,  it  has  been  thought  that  the 
preservation  depended  on  this  complete  elimination  of 
the  oxygen  at  100°  C.  (212''  P\). 

The  total  absence  of  water  very  efficaciously  opposes 
the  development  of  living  organisms.  Thus  we  can 
preserve,  as  we  may  say,  indefinitely,  dried  meat  and 
vegetables. 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  221 


AH  substances  known  as  antiseptics  are  also  enemies 
to  ferments.  Thus  common  marine  salt,  alcohol,  creo- 
sote, phenol,  salicylic  acid,  sulphurous  acid,  the  sul- 
phates, potassium  acetate  (Sacc),  carbon  dioxide,  tannin, 
the  acids,  many  metallic  salts,  as  those  of  copper,  mer- 
cury, iron,  aluminum  ;  potassium  chromates,  arsenious 
acid,  prussic  acid,  lime  water,  the  antiseptic  properties 
of  which  are  well  known  and  have  been  frequently  tried, 
all  these  are  also  poisons  for  ferments  of  various  kinds 
in  the  quantities  in  which  they  are  active. 

The  preservative  action  of  oil,  grease,  ashes,  fine  sand, 
bran,  sawdust,  coatings  of  paraffin  or  gelatine,  is  explained 
by  these  porous  or  impermeable  bodies  preventing  the 
approach  and  access  of  germs  brought  by  the  air,  like 
the  cotton  wool  in  Schroeder's  experiment. 

Products  of  Putrefaction. — The  products  of  putrefac- 
tion are  very  numerous.  This  may  be  easily  under- 
stood, first,  because  the  putrid  change  of  an  organ  or 
liquid  directly  taken  from  the  animal  or  vegetable 
economy  is  the  resultant  of  the  decomposition  of  the 
various  constituents  which  are  found  in  it.  The  special 
study  of  the  products  of  putrefaction  of  each  particular 
albuminoid  substance  has  only  been  attempted  in  a  very 
few  cases. 

In  the  second  place,  the  compounds,  definite  in  appear- 
ance, which  undergo  putrid  fermentation,  are  so  complex 
in  their  constitution,  that  we  ought  to  expect  to  meet 
with  a  great  number  of  derivatives  formed  by  putrid 
decomposition. 

The  most  constant  products  which  make  their  appear- 
ance in  putrefactions  screened  from  the  air  are  leucine, 
and  probably  some  of  it  homologues,  tyrosine,  the  vola- 


222 


ON  FERMENTATION. 


tile  fatty  acids  of  the  series  C°  H^n  (formic,  acetic, 
propionic,  butyric,  valerianic,  caproic,  &c.),  ammonia,  and 
some  compound  ammonias  (ethylamine,  propylamine, 
amylamine,  trimethylamine),  carbon  dioxide,  sulphu- 
retted hydrogen,  hydrogen,  and  nitrogen. 

If  we  refer  to  what  will  be  presently  said  with  refer- 
ence to  the  decomposition  of  albuminoid  substances 
under  the  influence  of  barium  hydrate,  we  shall  be  able 
easily  to  account  for  the  appearance  of  these  various 
products.  On  one  hand,  the  albuminoids  contain  the 
elements  of  urea,  and  ought  to  be  considered  as  com- 
pound ureids.  This  fact  alone  explains  the  appearance 
of  carbon  dioxide,  and  of  a  part  of  the  ammonia.  {See 
ammoniacal  fermentation.) 

The  albuminoids  are  decomposed  by  hydratation 
under  the  influence  of  baryta,  furnishing  leucine  and 
some  of  its  homologues,  tyrosine  and  a  sulphide.  These 
first  products  may,  probably,  undergo  the  ulterior  action 
of  ferments,  and  yield  ammonia  and  volatile  fatty  acids. 
We  know,  in  fact,  that  in  presence  of  putrefied  fibrin, 
leucine  is  resolved  into  ammonia  and  valerianic  acid. 

O  H^^  N  02  -i-  2      O  =  O^  -h  N       +  CO^  + 

Everything  leads  us  to  believe  that  putrefaction  is  a 
complex  phenomenon — that  it  is  only  a  successive  series 
of  fermentations  exerted  on  more  and  more  simple 
products. 

Thus,  for  example,  when  we  leave  fibrin  to  sponta- 
neous decomposition,  without  access  of  air,  it  is  resolved 
into  two  principles,  as  under  the  influence  of  sea- 
salt.  One  of  these  principles  is  albumin,  which,  on 
account  of  its  greater  resistance  to  the  action  of  fer- 


DUTYRIC  FERMENTATION  AND  PUTREFACTION.  223 

ments,  will  be  found  for  a  long  time  in  the  putrid  liquid. 
The  second  product  of  this  decomposition,  undergoing 
somewhat  quickly  a  more  thorough  change,  yields  acetic, 
butyric,  valerianic,  and  capric  acids,  as  well  as  ammonia 
(Brendecke),  which  are  evidently  derived  from  the 
amido-acids  homologous  with  leucine. 

The  chemical  reactions  which  accompany  the  putre- 
faction of  the  albuminoids  are,  then,  for  the  most  part, 
phenomena  of  hydratation,  which  may  be  reproduced 
identically  by  chemical  forces  alone,  independently  of 
vital  action.  Thus  we  shall  see  that  phenomena  of  this 
kind  may  be  excited  by  the  action  of  soluble  ferments, 
whether  diastasic  or  indirect ;  and  we  are  induced  to 
suppose  that  a  part,  at  least,  of  the  transformations 
undergone  by  proteids,  and  their  more  immediate  deriva- 
tives, are  the  consequences  of  phenomena  of  this  order 
(indirect  fermentation). 

Nothing  resembles  putrid  fermentation,  with  reference 
to  the  derived  products,  more  nearly  than  the  change 
which  takes  place  in  the  constituent  parts  of  yeast, 
when  left  to  itself  without  nourishment,  deprived  of  sugar 
and  oxygen. 

We  see,  in  fact,  the  appearance  of  leucine,  tyrosine, 
sarcine,  &c.  This  is  the  first  step  ;  the  action  stops 
there,  and  goes  no  farther ;  the  yeast,  or  the  special 
soluble  ferment  which  it  secretes,  is  unfit  to  attack 
these  bodies  again  ;  but  if  we  wait  for  the  development 
of  vibrios,  we  shall  find  the  production  of  ammonia, 
carbon  dioxide,  and  volatile  fatty  acids,  at  the  same 
time  that  the  leucine  partly  disappears. 

M.  Ulysse  Gayon  has  published  quite  recently,  as 
a  thesis  for  the  *'Doctorat  es  Sciences"  (Paris,  1875, 


224 


ON  FERMENTATION. 


Faculte  des  Sciences  de  Paris,  No.  362),  the  result  of 
many  experiments  on  the  spontaneous  decomposition  of 
eggs.  The  question  was  important,  and  very  interesting 
to  the  adversaries  of  the  theory  of  spontaneous  genera- 
tion. Besides,  the  facts  observed  by  M.  Donne  and 
M.  Bechamp  on  this  subject  seemed  contrary  to  the 
ideas  of  M.  Pasteur  on  the  general  cause  of  putre- 
faction. M.  Gayon,  a  pupil  and  demonstrator  of 
M.  Pasteur's,  endeavoured  to  bring  the  spontaneous 
decomposition  of  eggs  and  their  putrefaction  under  the 
general  law  enunciated  by  his  teacher. 

M.  Donne  (Experiences  sur  I'Alteration  spontanee 
des  CEufs,  Comp.  Rend,  de  I'Ac.  57,  p.  450,  1863)  had 
said  :  "  If  we  take  eggs  in  their  natural  state,  not  shaken, 
and  leave  them  to  themselves,  they  remain  for  weeks 
and  months,  even  during  the  great  heat  of  summer, 
without  undergoing  any  putrid  decomposition.  The 
egg  has  no  unpleasant  smell,  and  nothing,  either  pos- 
sessing animal  or  vegetable  life,  is  produced,  either  on 
the  surface  of  the  membrane  or  in  the  inside  ;  there 
are  no  traces  of  infusoria  or  microscopical  vegetation. 

"  If,  on  the  contrary,  we  destroy  the  physical  struc- 
ture of  the  interior  of  the  egg  by  shaking ;  if,  that  is 
to  say,  we  break  up  the  texture,  and  the  cells  of  the 
albuminous  substance,  and  thus  mix  together  the  yolk 
and  the  white,  then,  even  without  access  of  the  external 
air,  and  even  guarding  against  this  intervention  by  extra 
precautions,  such  as  a  coating  of  collodion  spread  over 
the  surface  of  the  egg,  we  find  all  the  phenomena  of 
decomposition  make  their  appearance,  after  a  longer  or 
shorter  time,  according  to  the  temperature,  but  always 
in  less  than  a  month  ;  all  the  phenomena  of  decomposi- 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  225 

tion,  with  the  exception,  however,  of  the  production  of 
living  organisms,  either  vegetable  or  animal ;  for,  what- 
ever may  be  the  degree  of  rottenness  to  which  we  allow 
the  egg  to  proceed,  we  can  never  discover  the  slightest 
trace  of  animalculae,  or  of  microscopic  vegetable  life ; 
the  matter  of  the  egg  grows  troubled,  and  of  a  livid 
colour ;  it  exhales  a  fetid  odour  directly  we  break  the 
shell,  but  nothing,  absolutely  nothing,  stirs  in  its  sub- 
stance ;  nothing .  lives,  and  the  most  careful  and  fre- 
quently repeated  examination  by  means  of  the  micro- 
scope does  not  enable  us  to  discover  the  least  trace  of 
an  organized  or  living  being.'* 

We  may  add,  that  M.  Bechamp  also  found  no  orga- 
nisms in  rotten  eggs. 

M.  U.  Gayon's  experiments,  into  the  details  of  which 
we  cannot  enter,  led  him  to  the  following  conclusions  : — 

"  Putrefaction  in  eggs,  whether  in  the  presence  or  the 
absence  of  air,  is  correlative  to  the  development  and 
multiplication  of  microscopical  organisms  of  the  family 
of  vibriones, 

"  In  other  terms,  contrary  to  the  result  found  by  M. 
Donne  and  M.  Bechamp,  eggs  make  no  exception  to 
the  great  law  of  correlation  which  M.  Pasteur  has 
demonstrated  for  all  the  phenomena  of  fermentation, 
properly  so  called.'' 

We  are  thus,  on  the  subject  of  eggs,  confronted  by 
two  distinct  affirmations,  as  much  opposed  as  black 
and  white.  M.  Donne  found  them ;  M.  Gayon  did 
not.  We  have  no  balance  wherewith  to  estimate  and 
compare  the  skill  of  the  two  observers.  It  appears  to 
us  certain  that  M.  Gayon  saw  what  he  described  ; 
but  we  cannot  affirm  that  M.  Donne  was  absolutely 


226 


ON  FERMENTATION. 


mistaken,  and  that  the  eggs,  in  the  conditions  under 
which  he  placed  them,  contained  vibrios  which  he  did 
not  find. 

In  the  absence  of  any  other  criterion,  we  bring  for- 
ward a  very  important  fact,  mentioned  by  M.  Gayon 
himself. 

This  skilful  microscopist  observed  that  some  of  the 
eggs  experimented  upon  at  the  temperature  of  about 
25°  C.  (77°  F.)y  whether  shaken  or  no,  underwent  a 
special  modification,  distinct  from  ordinary  putridity 
and  from  acid  fermentation.*- 

The  decomposed  mass  is  of  a  dirty  yellow  colour, 
it  has  an  odour  of  dried  animal  matter,  and  is  very  fluid  ; 
we  see  in  it,  also,  a  great  number  of  needle-like  as  well 
as  botryoidal  crystals,  formed  of  tyrosine.  It  contains 
much  greater  quantities  of  tyrosine  and  leucine  than 
are  found  in  ordinary  putrefaction.  M.  Gayon  was  not 
able  to  discover,  under  these  circumstances,  any  trace 
of  microscopic  organisms,  either  in  the  inside,  on  the 
surface,  or  in  the  substance  of  the  membranes.  How- 
ever, tyrosine  and  leucine  are  evident  and  unquestion- 
able symptoms  of  the  decomposition  of  albuminoid 
matter. 

Between  the  production  of  these  substances  and  the 

*  In  certain  cases,  M.  Gayon  saw  the  contents  of  eggs,  especially  of  those 
which  had  been  shaken,  transformed  into  a  homogeneous  mass,  of  the  con- 
sistence of  butter,  and  of  a  bright  yellow  colour,  with  a  sour  smell,  and  a 
strongly  acid  reaction.  M.  Bechamp  observed  a  similar  change  in  an  ostrich 
egg,  and  was  able  to  distinguish  the  presence  of  alcohol,  acetic  acid,  and 
sulphuretted  hydrogen.  M.  Gayon  attributes  this  change,  to  which  he  gives 
the  name  of  acid  fermentation,  to  the  presence  of  special  organisms.  These 
are  small  immovable  rods,  with  pale  outlines  and  homogeneous  tints,  either 
isolated  or  articulated  two  and  two  together,  from  ^.-^^  to  of  a  millimbtrc 
in  length,  and  j-^^^jj  to  jy^^g  of  a  millimetre  in  thickness. 


BUTYRIC  FERMENTATION  AND  PUTREFACTION.  22/ 


phenomena  called  putrefaction  there  is,  chemically 
speaking,  no  very  clear  distinction  to  be  drawn.  They 
are  reactions  of  the  same  order,  decompositions,  more 
or  less  extensive,  of  the  proteid  molecule ;  the  traces 
of  sulphuretted  hydrogen,  and  other  fetid  products 
which  communicate  such  a  repulsive  odour  to  putrefac- 
tion, cannot  serve  to  establish  an  absolute  and  philoso- 
phical line  of  demarcation  between  the  decomposition 
without  organisms  observed  by  M.  Gayon,  and  what  is 
wrongly  termed  putrefaction  properly  so  called. 

The  result  of  this  seems  to  be,  that  albuminoid  mat- 
ters are  able  to  undergo  certain  decompositions,  certain 
changes,  without  the  intervention  of  living  organisms. 

By  means  of  a  very  simple  and  ingenious  apparatus, 
M.  Gayon  succeeded  in  extracting  the  gas  contained 
in  large  ostrich  eggs  in  a  state  of  putrefaction. 

One  of  these  eggs,  in  a  state  of  thorough  putrefaction, 
yielded  150  cubic  centimetres  of  gas,  containing  per 
cent : — 

Sulphuretted  hydrogen 
Carbon  dioxide  . 
Hydrogen    .  . 
Nitrogen  . 

lOO'O 


Traces. 

30-5 
40*2 

29'3 


The  presence  of  nitrogen  might  be  due  to  the  accumula- 
tion of  a  certain  quantity  of  air  in  the  air-bubble  before 
putrefaction. 

Among  the  solid  and  liquid  products  of  the  putrefac- 
tion of  eggs,  the  presence  of  small  quantitiesof  leucine  and 
tyrosine,  alcoholic  products,  and  volatile  acids  (butyric 
acid)  were  recognized.    The  sugar  had  disappeared. 


228 


ON  FERMENTATION. 


CHAPTER  XII. 

FERMENTATION  BY  OXIDATION. 

Acetic  fermentation  and  the  reactions  which  we  shall 
class  with  it  under  the  generic  name  of  fermentation  by 
oxidation,  have  a  special  character,  which  we  have 
not  met  with  in  any  of  the  phenomena  which  we  have 
hitherto  studied. 

Not  only  are  the  fermentable  matter  and  the  ferment 
concerned  in  the  reaction,  the  one  furnishing  the  con- 
stituent parts  of  the  new  bodies  which  are  formed,  and 
the  other  acting  as  a  cause,  but  we  find  that  a  third 
factor,  the  oxygen  of  the  air,  becomes  necessary. 

In  other  words,  under  the  name  of  fermentation  by 
oxidation,  we  shall  speak  of  combustions  set  up  by 
living  organisms,  which  serve  as  media  between  the 
oxygen  of  the  air  and  the  combustible  body  or  ferment- 
able matter.  As  to  the  results  of  this  combustion,  they 
may  vary,  according  to  the  nature  of  the  body  which 
is  burnt,  and  may  even  be  resolved  into  the  simple  pro- 
ducts of  the  most  complete  combustion  (water  and 
carbon  dioxide). 

We  will  begin  with  the  acetic  fermentation  of  alcohoL 
It  has  been  long  known  that  the  alcohol  contained  in 
fermented  liquids,  such  as  wine,  beer,  &c.,  will  disappear 
under  certain  circumstances,  and  give  place  to  vinegar 


FERMENTATION  BY  OXIDATION.  229 

or  acetic  acid,  and  that  the  air,  or  rather  its  oxygen, 
plays  a  part  in  this  reaction.  The  progress  of  chemistry, 
and  the  exact  determination  of  the  respective  composi- 
tion of  alcohol  and  acetic  acid,  give  a  simple  .and  clear 
account  of  the  reaction  ;  it  may  be  formulated  thus  : — 

O  H6  0  +  02  =      O  +  C2  H4  O^ 

Alcohol.  Acetic  acid. 

The  oxidation  may  take  place  by  two  reactions,  with 
the  production  of  an  intermediate  product,  aldehyde  : — 

H6  O  +  O  =  H2  O  -f  C2  H4  O 
Alcohol.  Aldehyde. 
C2      O  +  O  =  C2  H4  02 
Aldehyde.  Acetic  acid. 

In  a  purely  chemical  point  of  view,  we  have  here  a 
very  simple  phenomenon,  on  which  we  need  not  dwell. 

The  determining  causes  of  this  oxidation  will  have 
greater  claim  on  our  attention.  Dobereiner  having 
shown  by  an  experiment,  which  has  become  classic,  that 
the  vapour  of  alcohol  mixed  with  the  oxygen  of  the 
air  becomes  acid,  being  transformed  into  acetic  acid, 
thought  that  he  had  ascertained  by  this  means  the  true 
theory  of  the  phenomenon,  the  essential  conditions  of 
which  became  the  simultaneous  action  of  alcohol  and 
oxygen,  in  presence  of  a  porous  body,  such  as  finely 
divided  platinum,  charcoal,  wood  shavings,  &c.,  capable 
of  favouring  by  a  catalytic  action,  that  of  contact,  the 
oxidation  of  the  alcohol. 

It  was  on  this  idea  that  the  process  of  rapid  acetifica- 
tion,  called  the  German  process,  was  founded  ;  this  was 
first  tried  by  Schiitzenbach  in  1823.  Long  before  this, 
Boerhaw  had  already  introduced  into  practice  an  analo- 


230 


ON  FERMENTATION. 


gous  commercial  process.  He  employed  vats  three 
metres  high  and  one  and  a  half  metres  in  diameter,  with 
a  double  bottom  pierced  with  holes,  placed  at  30  centi- 
metres (about  a  foot)  from  the  bottom.  Bunches  of 
grapes,  from  which  the  juice  had  been  expressed,  were 
placed  on  this  false  bottom,  so  as  to  fill  the  vat.  One  of 
the  vats  was  entirely  filled  with  wine,  the  other  was  only 
half  filled  ;  after  twenty-four  hours,  liquor  was  drawn  from 
the  full  vat  and  poured  into  the  other  so  as  to  fill  it ;  this 
operation  was  alternately  repeated  till  the  acidification 
was  complete.  It  is  seen  that,  in  the  vat  which  is  half 
full,  the  alcoholic  liquor  which  soaks  the  grape  stalks 
and  skins  is  exposed  to  the  action  of  air  on  a  large 
surface,  and  the  oxidation  is  singularly  favoured  by  this 
fact. 

In  Schiitzenbach's  process,  use  is  made  of  a  large  oak 
vat,  from  two  to  three  metres  in  height  by  one  in  diam- 
eter, furnished  with  a  false  bottom  pierced  with  holes, 
and  placed  about  30  centimetres  from  the  bottom. 
Some  centimetres  higher,  the  circumference  of  the  vat  is 
regularly  pierced  with  a  series  of  holes  passing  entirely 
round  it.  These  orifices  are  inclined  from  without 
inwards,  so  as  to  prevent  the  liquor  from  escaping. 

At  the  upper  part,  at  the  distance  of  30  centimetres 
from  the  lid,  another  false  bottom  is  placed,  pierced  with 
many  small  holes  and  several  large  ones  ;  the  latter 
are  usually  closed  by  plugs,  and  are  intended  for  the 
purpose  of  renewing  the  air,  when  it  is  deoxygenated. 
The  whole  is  closed  by  a  cover  furnished  with  an  open- 
ing carrying  a  funnel  which  can  be  closed,  and  serves 
for  the  introduction  of  the  liquor  to  be  oxidated.  The 
interval  between  the  two  partitions  is  filled  with  beech- 


FERMENTATION  BY  OXIDATION. 


231 


shavings.  A  thermometer  placed  in  the  interior  of  the 
vat  gives  an  idea  of  the  intensity  of  the  reactions. 

All  being  thus  arranged,  hot  vinegar  is  first  poured 
into  the  vat ;  this  filters  through  the  shavings,  impreg- 
nates them,  and  serves  afterwards  to  facilitate,  or  rather 
to  set  up,  the  oxidation  of  the  alcohol.  In  order  to  be 
transformed  into  vinegar,  suitable  mixtures  of  alcohol 
and  vinegar  are  employed,  similar  to  those  which  are 
used  in  the  Orleans  process.  The  temperature  of  the 
surrounding  air  is  maintained  at  21°  C.  (about  70°  F.). 
The  alcoholic  liquor  employed  is  heated  to  between  26° 
and  27°  C.  (79°  and  81°  F.) ;  the  temperature  rises  spon- 
taneously to  38°  to  42°  C.  (101°  or  108°  F.)  in  the  vats. 

The  alcoholic  liquor  is  never  completely  oxidized  by 
its  first  passage  through  the  vat ;  the  operation  is  once 
or  twice  repeated,  either  in  the  same  vat,  or  in  others  by 
the  side  of  it.  (For  further  details  see  special  works  on 
chemical  technology.)  In  good  manufactories,  a  result 
is  obtained  which  does  not  differ  by  more  than  six  per 
cent,  from  that  indicated  by  theory ;  and  even  this  loss 
may  be  lessened  by  certain  suitable  accessory  arrange- 
ments, to  which  we  need  not  now  allude. 

The  French  method  of  acidification  of  wine,  called 
the  Orleans  method,  is  very  different.  We  will  say  a 
few  words  about  it  before  we  enter  on  the  results  of 
Pasteur's  researches,  and  the  commercial  consequences 
which  have  been  derived  from  them. 

The  oxidation  takes  place  in  barrels  placed  side  by 
side,  on  wooden  frames,  supported  by  stone  pillars.  At 
the  upper  front  part  of  each  vat  two  orifices  of  unequal 
size  are  made  ;  the  larger  serves  for  the  purpose  of 
introducing  and  drawing  off"  liquor,  the  other  for  the 


232 


ON  FERMENTATION. 


admission  of  air.  These  vats,  which  contain  from  two 
to  four  hundred  Htres  (44  to  88  gallons)  called  mothers, 
are  at  first  one-third  filled  with  strong  boiling  vinegar  ; 
from  II  to  12  litres  (20  to  21  pints)  of  wine  are  then 
added,  and  the  vats  are  then  left  undisturbed  ;  at  the 
end  of  a  week's  acetification,  another  quantity  of  wine 
is  added,  and  so  on  from  time  to  time  till  the  vessel  is 
half  full  of  vinegar.  A  third  part  of  the  contents  of 
the  "  mother"  is  then  drawn  out  by  means  of  a  siphon, 
and  wine  is  again  added  from  time  to  time  in  portions 
of  II  to  12  litres  (20  or  21  pints).  A  regular  and 
continued  process  is  thus  carried  on.  It  is  often  found 
that,  without  any  apparent  cause,  a  certain  "  mother " 
refuses  to  acidify  the  wine,  or  only  gives  rise  to  a  very 
slow  and  tedious  oxidation ;  in  this  case,  the  employ- 
ment of  a  much  stronger  wine,  or  a  much  higher  temper- 
ature, sometimes  restores  the  action  to  its  normal  activity. 
These  anomalies  have  only  been  explained  by  Pasteur  s 
experiments.  The  operation  in  one  barrel  is  considered 
to  be  terminated  when  a  stick  plunged  into  the  liquid  is 
covered  with  a  thick  white  froth  (flour  of  vinegar)  ;  as 
long  as  the  froth  is  red,  the  addition  of  wine  is  continued. 
The  most  favourable  temperature  is  between  24°  and 
27°  C.  (76°  to  82°  R). 

In  this  special  manufacture  of  vinegar,  the  influence 
of  porous  bodies  in  determining  the  oxidation  cannot 
be  brought  to  bear,  as  in  the  experiment  with  platinum 
or  in  the  German  method  of  manufacture.  The  vinegar- 
makers,  from  their  special  experience,  attribute  the 
acidification  to  the  action  of  a  deposit  which  forms 
in  the  barrels,  and  to  which  they  give  the  name  of 
„  mother  of  vinegar." 


FERMENTATION  BY  OXIDATION.  233 

According  to  the  opinions  of  Liebig,  which  have  so 
long  prevailed  in  scientific  matters,  either  dead  or  living 
organic  matter,  when  in  contact  with  alcohol  in  wine, 
possess  the  property,  after  having  absorbed  oxygen,  of 
oxidizing,  at  the  ordinary  temperature,  organic  and  inor- 
ganic substances.  This  property  would  therefore  exist 
in  solid  organic  substances  which  are  in  a  state  of  decom- 
position or  putrefaction.  To  support  this  view,  Liebig 
(Ann.  de  Chim.  (4),  vol.  23,  p.  178,)  recalls  de  Saussure's 
experiment,  who  found  that  soil  placed  in  a  mixture  of 
hydrogen  and  oxygen  gave  rise  to  the  formation  of 
water  and  the  disappearance  of  hydrogen,  and  of  an 
equivalent  quantity  of  oxygen.  We  may  state  in  a  few 
words,  and  without  entering  into  too  many  details  on  a 
question  which  seems  determined,  that  Liebig  explains 
the  production  of  acetic  acid  by  a  certain  movement 
communicated  by  principles  in  process  of  decomposition 
— a  movement  which  in  this  case  excites  oxidation. 

In  fact,  his  ideas  are  wanting  in  clearness.  Sometimes 
he  seeks  for  the  key  to  the  phenomenon  in  a  catalytic 
action  of  porous  bodies ;  sometimes,  with  reference  to 
his  general  theory  of  fermentation,  he  attributes  it  to  the 
influence  of  a  ferment  (organic  matter  in  process  of 
decomposition). 

Such  was  the  state  of  the  question  when  Pasteur 
undertook  his  researches  on  the  causes  of  the  sponta- 
neous acidification  of  wine  and  alcoholic  liquors. 

According  to  him,  the  oxidation  of  alcohol  is  the  con- 
sequence of  the  action  of  a  cryptogam  of  the  genus 
Mycoderma,  We  will  give  a  summary  of  the  experi- 
ments on  which  his  opinion  was  founded. 

If  on  the  surface  of  any  organic  liquid,  necessarily 
11 


234  ON  FERMENTATION. 

containing  phosphates  and  nitrogenous  organic  matter, 
we  allow  any  species  of  mycoderma  to  develop  itself, 
until  the  whole  surface  of  the  liquid  is  covered  with  it ; 
if  then  we  carefully  remove  the  nutritive  liquid  by  means 
of  a  siphon,  without  suffering  any  portion  of  the  mem- 
branes to  break  up  ;  then,  if  we  substitute  for  this  liquid 
an  equal  volume  of  water,  containing  lo  per  cent,  of 
alcohol,  we  immediately  see  the  plant  which  is  placed 
under  these  abnormal  circumstances  of  nutrition,  set  up 
a  reaction  between  the  oxygen  of  the  air  and  the  alcohol 
of  the  liquid.  The  acetification  begins  immediately, 
and  goes  on  with  great  activity.  After  a  certain  time, 
the  action,  impeded  by  the  great  acidity  of  the  liquid, 
proceeds  more  slowly ;  but  we  can  restore  to  it  all  its 
activity  by  substituting  alcoholized  water  for  the  acid 
liquid.  A  time,  however,  comes  when  the  plant,  becom- 
ing partly  decomposed  itself,  communicates  to  the  liquid, 
in  consequence  of  the  organic  and  mineral  ele*ments  of 
its  dead  tissues,  properties  which  serve  as  nutriment  for 
the  various  species  of  mycoderma.  The  action  then  takes 
on  a  different  phase  ;  the  acetic  acid  and  alcohol  disap- 
pear with  ^reat  rapidity,  and  the  liquid  becomes  com- 
pletely neutralized  ;  because  as  soon  as  the  plant  finds 
in  the  subjacent  medium  the  nutritive  principles  suited 
to  its  development,  it  sets  up  much  more  intense  oxi- 
dizing action,  and  burns,  not  only  the  alcohol,  but  also 
the  acetic  acid,  converting  it  into  water  and  carbon 
dioxide. 

This  complete  combustion  is  noticed  whenever  we 
cause  the  mycodermata  to  be  developed  on  alcoholic 
liquids  containing  food  fit  for  the  nourishment  of  the 
plant,  such  as  v/inc,  beer,  or  fermented  organic  liquids  ; 


J 


FERMENTATION  BY  OXIDATION.  235 


unless,  however,  we  place  the  mycoderma,  whether  inten- 
tionally or  unintentionally,  under  the  conditions  of 
incomplete  or  tardy  development,  that  is  to  say,  in  a 
sickly  state. 

To  sum  up,  we  may  say  that  the  mycodermata,  devel- 
oping on  the  surface  of  an  alcoholic  liquid  which  con- 
tains suitable  nutritious  principles,  burn  the  alcohol,  and 
bring  it  to  the  same  state  as  oxygen  at  red  heat,  that  is, 
complete  combustion.  If,  on  the  contrary,  we  diminish 
the  vital  activity  of  the  mycoderma,  whether  by  depriv- 
ing it  of  its  nourishment  or  by  any  other  means,  the 
oxidizing  action  which  it  may  be  able  to  set  up  will  not 
go  so  far,  and  the  alcohol  may  change  into  acetic  acid. 
M.  Pasteur's  experiments  will  also  show  that  the 
influence  attributed  to  ordinary  porous  organized  bodies, 
in  the  German  manufacture  of  vinegar,  is  the  result  of 
imperfect  observation. 

The  beech-shavings  act,  not  on  account  of  their 
porosity,  but  because  their  surface  is  covered  with  thin 
pellicles  of  mycoderma  ;  the  many  points  of  contact 
with  the  air  favour  the  action,  but  are  not  the  deter- 
mining causes  of  it.  To  prove  this,  M.  Pasteur 
caused  some  alcohol  diluted  with  water  to  trickle  down 
a  cord.  The  drops  which  fell  from  the  end  of  the  cord 
did  not  contain  the  smallest  quantity  of  acetic  acid. 
The  experiment  lasted  for  more  than  a  month,  the 
liquid  trickling  extremely  slowly,  only  one  drop  in  two 
or  three  minutes.  If  we  repeat  this  experiment,  having 
previously  steeped  the  cord  in  a  liquid  on  the  surface  of 
which  there  is  a  pellicle  of  mycoderma,  a  portion  of 
which  clings  to  the  cord  as  it  is  withdrawn,  the  alcohol, 
which  passes  slowly  down  this  cord,  will  be  charged  with 


236 


ON  FERMENTATION. 


acetic  acid,  and  this  acetification  may  be  continued  for 
several  weeks. 

M.  Mayer  made  some  similar  experiments,  which 
entirely  confirmed  Pasteur's  results.  Thus  filter  paper, 
previously  boiled  with  hydrochloric  acid  at  50  per  cent., 
then  with  caustic  soda  at  50  per  cent,  and  finally  washed 
with  distilled  water,  was  laid  on  the  surface  of  an  alco- 
holic liquid,  without  giving  rise,  even  after  the  expiration 
of  a  month,  to  the  least  trace  of  acetic  acid.  The  result 
was  also  as  negative  when  a  funnel  was  filled  with  frag- 
ments of  glass  and  of  the  same  paper,  and  alcohol  at 
9  per  cent,  was  allowed  to  run  slowly  through  this  filter- 
ing mass. 

M.  Pasteur  did  not  go  more  deeply  into  this  ques- 
tion, and  did  not  ascertain  by  what  means  the  mycoder- 
mata  could  thus  give  rise  to  more  or  less  energetic 
oxidations. 

In  one  of  his  latest  writings  on  this  question  (Reponse 
aux  Critiques  de  M.  Liebig,  Ann.  Chim.  Phys.  (4), 
vol.  25,  p.  148,  1872),  he  thus  expresses  himself: — 

**This  little  microscopical  plant  {Mycoderma  aceti) 
possesses  the  power  of  condensing  the  oxygen  of  the 
air  in  the  same  manner  as  spongy  platinum  or  blood- 
globules,*  and  of  conveying  this  oxygen  to  matter 
beneath.'* 

It  seems,  from  these  few  words,  that  M.  Pasteur 
compares  the  action  of  his  organic  ferment,  the  Myco- 
dcnna  aceti,  to  that  of  spongy  platinum :  his  theory  of 
acetic  fermentation  really  differs  very  slightly  from  that 

*  The  oxygen  of  the  globules  of  blood  is  fixed  in  the  haemaglobin,  and  its 
oxidizing  power  is  scarcely  more  energetic  than  that  of  ordinary  dissolved 
oxygen. 


FERMENTATION  BY  OXIDATION.  237 


of  Liebig.  The  latter  admits  that  inanimate  porous 
substances  participate  in  the  properties  of  spongy  pla- 
tinum ;  Pasteur,  on  the  contrary,  attributes  this  quality 
only  to  living  organisms. 

Mayer  s  experiments  tend  to  prove  that  oxidation  by 
mycodermata  is  a  special  biological  action,  which  cannot 
be  attributed  solely  to  the  physical  condition  of  the  plant 
which  acts  as  ferment.  In  fact,  we  have  only  to  heat  an 
alcoholic  liquid,  covered  with  its  pellicle  of  acetic  myco- 
derma  in  full  process  of  acidification,  and  we  shall  arrest 
all  oxidation  ;  yet  it  is  difficult  to  believe  that  under 
these  conditions  the  physical  state  has  been  sensibly 
modified. 

M.  Mayer  also  noticed  striking  differences  between 
the  mode  of  action  of  spongy  platinum  and  that  of  the 
Mycorderma  aceti.  For  the  first,  an  elevation  of  tem- 
perature above  35°  C.  (95°  F.)  favours  the  oxidation,  by 
augmenting  the  tension  of  the  vapour  of  alcohol,  while 
the  maximum  activity  of  the  Mycoderma  aceti  is  placed 
between  20°  and  30°  C.  (68°  and  86°  F.) ;  its  oxidizing 
power  is  destroyed  under  10°  C.  (50°  F.)  and  above' 
35°  C.  (95°  F.).  We  can  oxidize  very  concentrated 
alcohol  with  platinum ;  the  concentration  is  even  a 
favourable  factor  ;  while  for  physiological  acetification, 
the  alcohol  employed  ought  scarcely  to  contain  more 
than  10  per  cent,  of  ethyl  hydrate. 

The  immediate  and  practical  consequence  of  the 
results  obtained  by  Pasteur  is,  that  in  order  to  act 
efficaciously,  the  mycoderma  ought  to  be  in  contact, 
at  the  same  time,  with  the  air  and  the  alcoholic 
medium.  The  new  commercial  method  for  the  acetifi- 
cation of  fermented  liquids,  now  employed  to  some 


238 


ON  FERMENTATION. 


extent  at  Orleans,  is  founded  on  this  principle.  The 
following  is  the  process  : — 

The  Mycorderma  aceti  is  first  sowed  on  the  surface 
of  an  aqueous  liquid  containing  2  per  cent,  of  alcohol, 
I  per  cent,  of  vinegar,  and  traces  of  alkaline  and 
alkalin-earthy  phosphates.  When  the  surface  is 
covered  with  the  membrane,  the  alcohol  begins  to 
acidify.  This  action  being  fully  set  up,  some  alcohol, 
wine,  or  beer  mixed  with  alcohol,  is  added  every  day  to 
the  liquid,  in  small  quantities ;  this  is  continued  till  the 
oxidation  becomes  slower ;  the  acetification  is  then 
allowed  to  terminate,  and  the  vinegar  is  drawn  off. 
The  membrane  is  collected,  washed,  and  employed  for 
a  new  operation.  It  is  better  always  to  give  the  plant 
sufficient  alcohol,  so  that  its  activity  should  not  be 
exerted  on  the  acetic  acid.  Nor  ought  it  to  remain  too 
long  out  of  the  liquid,  or  it  would  lose  its  active  force ; 
finally,  it  is  better  to  moderate  its  development,  to  pre- 
vent burning  oxidation. 

A  vat,  one  square  metre  in  section,  and  containing 
from  50  to  100  litres  (from  11  to  22  gallons),  can  fur- 
nish per  day  from  5  to  6  litres  (9  to  10^  pints)  of 
vinegar.  The  successive  phases  of  the  operation  are 
ascertained  by  means  of  a  thermometer  divided  into 
tenths  of  a  degree,  the  bulb  of  which  is  plunged  into 
the  liquid. 

When  we  act  upon  diluted  alcohol,  it  is  better  to  add 
to  the  liquid  --^  ^^^^  of  a  mixture  of  magnesium  phos- 
phate, and  potassium  and  ammonium  phosphates. 

Special  arrangements  allow  the  introduction  of  the 
liquid,  without  there  being  any  necessity  for  disturbing 
the  superficial  film  of  mycodcrma.     The  vessels  are 


FERMENTATION  BY  OXIDATION. 


cylindrical,  or  of  a  prismatic  form,  of  one  square  metre 
in  section,  and  i  metre  in  depth,  closed  by  a  cover  fur- 
nished with  orifices  for  the  admission  of  the  air. 


Fig.  22. — Mycoderma  aceti. 

It  only  remains  for  us  to  say  a  few  words  about  the 
botanical  character  of  the  Mycorderma  aceti. 

The  continuous  membranes,  either  wrinkled  or 
smooth,  which  are  found  on  the  surface  of  liquids 
while  in  acetic  fermentation,  are  generally  formed 
(Fig.  22)  of  very  minute  elongated  cells,  whose  greater 
diameter  varies  from  i,  S,  to  3  thousandths  of  a  milh- 
metre  ('000059  to  'OOOiiS  in.)  ;  these  cells  are  united 
in  chains,  or  in  the  form  of  curved  rods.  Multiplica- 
tion seems  to  be  effected  by  the  transverse  division  of 
the  fully  developed  cells.  This  division  is  preceded  by 
a  median  constriction,  which  has  been  considered  by 
some  authors  as  a  morphological  characteristic  of 
the  cell. 

It  appears,  from  this  description,  that  the  myco- 
derma of  vinegar  belongs  to  the  family  of  bacteria. 

The  general  conditions  of  the  nutrition  of  acetic 
bacteria  have  been  discovered  by  Pasteur,  and  closely 
resemble,  up  to  a  certain  point,  those  of  beer-yeast. 

Thus  mineral  salts,  alkaline  and  alkalin-earthy  phos- 
phates, proteid  nitrogenous  substances,  or  ammoniacal 
salts,  are  elements  necessary  for  the  devolopment  of 


240 


ON  FERMENTATION. 


these  organisms.  Diluted  alcohol  (10  per  cent,  at  most) 
seems,  in  this  case,  to  take  the  place  of  hydrocarbon 
matter ;  it  may  be  supplemented  by  the  acetic  acid  ; 
for,  according  to  M.  Pasteur,  the  progressive  weakening 
which  vinegar  undergoes,  when  the  acetification  is  left 
too  long  to  itself,  is  due  to  a  subsequent  burning  of  the 
acetic  acid* 

M.  Blondeau  (Comp.  Rend.,  vol.  57,  p.  953)  has  even 
observed  that  sugar  can  acidify  without  passing  through 
the  condition  of  alcohol,  under  the  influence  of  the 
mother  of  vinegar. 

We  must,  however,  notice  that  the  activity  of  the 
ferment  is  increased  by  the  presence  in  the  liquid  of  a 
certain  quantity  of  acetic  acid. 

^  Antiseptic  agents,  in  general,  which  by  their  presence 
delay  arid  arrest  the  development  of  the  yeast  of  beer, 
and  consequently  alcoholic  fermentation,  act  in  the 
same  manner  with  respect  to  the  Mycoderma  aceti. 
Sulphurous  acid  is  especially  active  in  this  manner; 
and  it  is  partly  in  order  to  avoid  the  acetification  of 
wine,  that  care  is  taken  to  put  in  the  tuns  intended  to 
receive  it,  sulphurous  acid,  or  to  burn  sulphur  matches 
in  them. 

The  Mycoderma  vini^  of  which  we  have  spoken 
above  (p.  59),  resembles  in  many  respects  the  acetic 
ferment.  Like  it,  it  is  developed  on  the  surface  of 
fermented  alcoholic  liquids,  in  the  form  of  smooth  or 
wrinkled  films  or  membranes  ;  but  these  latter,  how- 
ever, are  thicker  and  more  compact.  It  acts  also  as 
the  means  for  conveying  the  oxygen  of  the  air  to 
the  alcohol  of  the  medium,  and  to  the  other  com- 
bustible principles  ;  but,  under  its  influence,  combustion 


FERMENTATION  BY  OXIDATION.  24 1 

is  complete,  and  accompanied  by  the  production  of 
carbon  dioxide  and  water  ;  it  is  to  this  action  that  we 
must  attribute  the  rapid  weakening  of  wines  covered 
with  mycoderma.  We  have  elsewhere  seen  that,  when 
immersed  in  the  midst  of  a  saccharine  liquid,  it  can 
act  like  the  yeast  of  beer,  and  produce  alcoholic  fer- 
mentation. 

Its  nutritive  principles  are  the  same  as  those  of  the 
mother  of  vinegar  (alcohol,  salts,  nitrogenous  com- 
pounds) ;  besides,  it  appears  also  capable  of  utilizing 
for  nutrition  certain  secondary  products  of  alcoholic 
fermentation,  such  as  succinic  acid  and  glycerin.  The 
forms  of  the  cells  of  this  mycoderma— forms  that  we 
know  to  be  variable,  seem  to  depend  in  a  great  degree 
on  the  conditions  of  nutrition. 

Its  activity  of  development  appears  included  between 
16^  and  30°  C.  (61°  and  86°  F.). 

Sloiv  ComkistioiL—ThQ  Mycodermata  of  which  we 
have  just  spoken  are  not  the  only  organized  ferments 
capable  of  exciting  the  slow  combustion  of  carburetted 
materials. 

It  has  long  been  known  that  organic  matter,  of  vege- 
table or  animal  origin,  left  in  contact  with  air,  undergoes 
progressive  and  complex  transformations,  known  under 
the  name  of  putrefaction,  of  slow  combustion,  and  of 
eremocausis,  whose  effect  is  to  transform  them  into 
principles  more  and  more  simple,  by  means  of  de- 
composition and  oxidation;  so  that,  in  the  end,  the 
carbon  is  restored  to  the  air  in  the  form  of  carbon 
dioxide,  the  hydrogen  under  the  form  of  water,  the 
nitrogen  either  as  free  nitrogen  or  ammonia.  M.  Pasteur 
has  distinguished,  among  the  complicated  facts  of  Dutrid 


242  ON  FERMENTATION. 

f  ermentation,  two  orders  of  distinct  phenomena,  although 
each  is  connected  with  the  reactions  set  up  by  hving 
organisms.  The  first  includes  the  putrefaction,  Avhich 
takes  place  without  the  assistance  of  oxygen  in  the  air, 
which  is  caused  by  the  presence  of  vibrios.  We  have 
spoken  of  this  in  the  chapter  on  butyric  fermentation, 
with  which  these  phenomena  are  connected. 

The  second,  slow  combustion,  is  due  to  bacteria, 
mucors,  and  mucidines,  that  is  to  say,  to  vegetable 
ferments,  which,  like  the  Mycoderma  viiii,  and  others, 
possesses  the  remarkable  property  of  exciting  the 
oxidation  of  a  great  number  of  organic  principles, 
such  as  sugars,  alcohols,  organic  acids,  albuminoid 
nitrogenous  matter,  &c.,  at  the  expense  of  the  oxygen 
of  the  air. 

After  having  proved,  by  careful  experiments,  to  which 
we  will  return  when  we  treat  on  the  origin  of  ferments, 
that  spontaneous  slow  combustion  of  animal  or  vegetable 
substances  depends  necessarily  on  the  development  of 
organisms  in  the  interior,  or  on  the  surface,  of  the 
substances  which  are  in  process  of  decomposition,  and 
that  without  organisms  there  is  neither  combustion  nor 
absorption  of  oxygen,  M.  Pasteur  traces  the  following 
picture  of  putrid  decomposition  in  contact  with  air 
(Comp.  Rend.,  June,  1863)  : — 

"  Even  the  most  easily  decomposed  animal  matter,  as, 
for  instance,  blood  or  urine,  may  be  preserved  for  an 
indefinite  length  of  time  in  air  which  has  been  calcined 
or  deprived  of  its  germs  ;  under  these  conditions,  the 
absorption  of  oxygen  is  but  trifling,  and  putrefaction 
does  not  take  place  ;  and,  at  the  same  time,  no  infusoria 
are  produced.    If,  on  the  contrary,  this  same  substance 


FERMENTATION  BY  OXIDATION.  243 


remains  exposed  to  the  ordinary  air,-  it  is  oxidized, 
putrefies,  and  infusoria  are  developed. 

"It  is  commonly  known  that  putrefaction  takes  a 
certain  time  to  declare  itself,  a  period  varying  according 
to  the  circumstances  of  temperature — of  the  neutral, 
acid,  or  alkaline  character  of  the  liquid.  Under  the 
most  favourable  circumstances,  at  least  twenty-four 
hours  are  required  before  the  phenomenon  begins  to 
manifest  itself  by  external  signs.  During  the  first 
period,  an  internal  movement  takes  place  in  the  liquid, 
the  effect  of  which  is  to  withdraw  entirely  the  oxygen  of 
the  air  which  is  in  solution,  and  to  substitute  for  it  carbon 
dioxide  gas.  The  total  disappearance  of  the  oxygen, 
when  the  medium  is  neutral  or  slightly  alkaline,  is  gene- 
rally due  to  the  development  of  the  smallest  kinds  of 
infusoria,  especially  the  Monas^  creptcscuhimy  and  the 
Bacterium  termo,  A  very  slight  troubling  then  takes 
place,  because  these  little  beings  pass  about  in  all  direc- 
tions. If  the  vessel  containing  the  putrescible  liquid 
has  a  large  opening  to  the  air,  the  bacteria  perish  only 
in  the  liquid  mass,  after  the  removal  of  the  oxygen, 
while  they  continue,  on  the  contrary,  to  propagate,  ad 
infinitum^  on  the  surface,  because  it  is  in  contact  with 
the  air.  There  they  cause  a  thin  film  to  form,  which 
goes  on  thickening  by  degrees,  until  it  falls  to  the  bottom 
of  the  vessel ;  then  another  forms,  and  so  on  continually. 
This  film,  to  which  different  mucors  and  mucidines  are 
attached,  prevents  the  solution  of  oxygen  gas  in  the 
liquid,  and  consequently  allows  the  development  of 
vibrios.  With  respect  to  these  latter  organisms,  the 
vessel  is  as  if  it  were  closed  against  the  introduction 
of  air. 


244 


ON  FERMENTATION. 


The  putrescible  liquid  thus  gives  rise  to  two  very 
distinct  kinds  of  chemical  action,  which  have  reference 
to  the  two  sorts  of  organisms  which  are  nourished  in  it. 
On  one  hand,  the  vibrios,  living  by  the  co-operation 
of  the  oxygen  of  the  air,  set  up  in  the  interior  of  the 
liquid  acts  of  fermentation — that  is  to  say,  they  transform 
the  nitrogenous  matter  into  more  simple,  but  still  com- 
plex, products. 

The  bacteria  (or  the  mucors),  on  the  other  hand,  con- 
sume these  same  products,  and  bring  them  to  the  state  of 
the  most  simple  ordinary  combinations,  water,  ammonia, 
and  carbon  dioxide. 

The  compounds  which  longest  resist  slow  combustion 
are  the  fixed  fatty  acids,  forming  the  adipocire  of  the 
vold  chemists,  cellulose  or  its  derivatives  formed  by 
dishydratation„  such  as  ulmic  acids,  vegetable  mould, 
peat,  &c. 

The  oleic  acid,  on  the  contrary,  disappears  altogether. 
But  little  is  known  of  the  details  of  these  various 
phenomena  of  slow  combustion. 

Can  the  same  organism  set  up  the  combustion  ot 
different  organic  principles,  differing  from  one  another 
in  their  constitution  }  Is  the  action  of  the  oxygen  pro- 
gressive, or  is  it  complete  from  the  very  commence- 
ment ?  These  and  many  other  secondary  questions, 
more  or  less  interesting,  present  themselves  ;  but  their 
solution  requires  long  and  minute  researches,  similar  to 
those  which  relate  to  alcoholic  fermentation.  However, 
the  cause  of  the  phenomena  is  known,  and  the  way  for 
new  investigations  is  open. 


CHAPTER  XIII. 


APPLICATIONS  OF  THE   RESEARCHES  AND    IDEAS  OF 
M.  PASTEUR. 

We  have  already  seen,  when  treating  of  acetic  fermen- 
tation, what  consequences  M.  Pasteur  has  drawn  from 
his  observations,  in  order  to  regulate  and  facilitate  the 
transformation  of  fermented  liquors  into  vinegar. 

The  manufacture  of  beer  may  also,  according  to  this 
investigator,  derive  advantage  from  the  careful  study  of 
fermentation  and  of  ferments. 

If  we  add  to  the  wort  of  beer  ordinary  yeast,  prin- 
cipally composed  of  cells  of  Saccharoinyces  cerevisice, 
with  very  few  foreign  organisms,  under  the  most  favour- 
able conditions  for  the  development  of  yeast,  it  will 
reproduce  almost  alone,  setting  up  direct  alcoholic  fer- 
mentation. As  the  quantity  of  original  yeast  is  found, 
after  the  fermentation,  to  be  six  or  seven  times  greater 
than  at  first,  supposing  that  no  foreign  germs  have  been 
introduced  from  without,  we  can  understand  that  the  new 
yeast  will  be  purer  than  the  first.  By  continuing  with 
this,  new  fermentations  of  the  wort  of  beer,  we  shall 
procure,  by  a  kind  of  selection  similar  to  that  described 
by  M.  Raulin,  with  reference  to  Aspergilltis  7iiger,  a  very 
pure  yeast,  free  from  the  mixture  of  other  organisms. 


246 


ON  FERMENTATION. 


When  this  result  has  once  been  obtained,  it  will  be  suffi- 
cient to  maintain  the  integrity  and  purity  of  ferment,  by 
excluding  from  contact  with  air  the  fermenting  vats  of 
beer,  and  by  carrying  on  the  process  in  closed  vessels, 
instead  of  open  vats.  The  principle  of  the  new  process 
patented  by  M.  Pasteur,  into  the  details  of  which  we 
cannot  enter,  depends  then  on  the  employment  of  pure 
yeast,  and  on  fermentation  with  the  exclusion  of  air,  in 
order  to  avoid  the  introduction  of  foreign  organisms, 
which,  by  their  ulterior  development,  might  produce 
changes  of  another  order,  lactic  fermentation,  &c. 

For  this  purpose,  the  wort,  after  it  is  prepared,  is 
drawn  off  while  boiling  into  vessels  made  of  wood  or 
metal,  and  cooled  in  a  current  of  carbon  dioxide, 
or  of  air  purified  from  ferments,  and  then  yeast  is 
added. 

The  beer,  after  the  first  fermentation,  is  drawn  off 
into  casks,  in  which  it  ripens  and  grows  clear.  The 
wort  may  be  carried  to  a  great  distance,  and  the  beer 
has,  according  to  M.  Pasteur,  superior  qualities  both  as 
to  taste  and  preservation. 

Preservation  of  Wine  —  M.  Pasteur  has  made  a  long 
and  very  attentive  study  of  the  changes  which  may  take 
place  in  wines  in  the  various  phases  of  their  preservation. 
His  observations  have  been  preserved  in  a  very  excellent 
work  published  on  this  subject.  We  can  only  give  here 
the  more  general  results  obtained  by  this  author.  He 
attributes  the  changes  which  take  place  in  wines  to  the 
development  of  special  living  ferments. 

A  special  organism  corresponds  to  each  order  of 
change,  i.e,,  to  each  disease  of  the  wine.  The  germs  of 
these  ferments  are  found  in  the  must  of  fermented  grapes, 


247 


248 


ON  FERMENTATION. 


and  may  develop  themselves  whenever  the  conditions 
become  favourable. 

In  order  to  prevent  their  development,  and  to  allow 
the  wine  to  be  kept  for  an  indefinite  term,  without  any 
change,  we  have  only  to  raise  the  temperature  for  a 
moment  to  60°  C.  (140°  F.).  The  germs  are  killed  at  this 
comparatively  low  temperature,  because  of  the  presence 
of  alcohol  (from  8  to  10  per  cent). 

The  warming  of  wines  for  the  purposes  of  preserving 
them  had  already  been  proposed  by  M.  Vergnette  de  la 
Motte,  but  the  rational  explanation  of  this  process  is 
due  to  M.  Pasteur.  He  also  determined  with  greater 
precision  the  conditions  of  the  operation. 

This  warming  may  be  effected  while  the  wine  is  in 
bottle  or  in  casks. 

Fig.  23  represents  the  apparatus  of  Giret  and  Vinas 
for  the  continuous  warming  of  wine.  It  is  composed  of 
a  stove  surmounted  by  upright  tubes  communicating  wdth 
the  chimney.  A  water-bath  completely  surrounds  these 
tubes.  The  cylinder  of  the  water-bath  is  fixed  to  the 
stove  by  means  of  two  rims,  between  which  is  a  slip  of 
cloth  saturated  with  paste.  The  two  rims  are  compressed 
by  iron  clamps,  so  that  this  cylinder  can  be  easily  re- 
moved. The  wine  circulates  in  a  cavity  formed  by  two 
concentric  cylinders,  secured  above  and  below  by  two 
annular  collars.  The  refrigerator  is  formed  of  a  cylinder 
containing  an  interior  cavity  similar  in  every  respect 
to  the  preceding  one.  The  cover  of  the  refrigerator 
is  movable,  and  is  fastened  by  an  arrangement  like 
that  which  serves  to  connect  the  cylinder  with  the 
stove. 

The  surfaces  in  contact  with  the  wine  are  tinned.  The 


RESEARCHES  OF  M.  PASTEUR. 


249 


water-bath  contains  water,  the  refrigerator,  as  well  as  the 
enclosure  around  it,  only  holds  wine. 

The  arrows  show  the  direction  of  the  circulation;  a 
thermometer  placed  within  the  bulb  of  the  tube  which 
connects  the  water-bath  with  the  refrigerator  gives  the 
maximum  temperature.  (For  further  details  on  this 
subject  see  La  Chimie  Technolique  de  Wagner,"  French 
translation.)* 

Application  of  M.  Pasteicr's  ideas  to  Pathology, — Some 
years  since  I  wrote  in  the  following  terms  (La  Chimie 
appliquee  a  la  physiologic  animale  a  la  pathologic  et  au 
diagnostic  medical,  par  P.  Schiitzenberger,  1864)  : — - 

All  diseases,  contagious  either  by  inoculation  or  by 
more  or  less  direct  contact,  whether  epidemic  or  endemic, 
are  evidently  produced  by  the  introduction  of  foreign 
poisonous  substances  into  the  living  organism,  producing 
true  poisoning.  When  in  an  affection  of  this  kind,  as  for 
example,  in  cholera,  yellow  fever,  or  malignant  ulcers, 
the  general  symptoms  and  the  mode  of  evolution  have 
a  well-marked  constancy  of  character,  notwithstanding 
differences  of  race,  species,  and  individuals,  one  is  com- 
pelled to  admit  the  specific  nature  of  the  poison  which 
gives  rise  to  certain  pathological  manifestations. 

"  These  conclusions,  drawn  from  many  facts,  observed 
in  every  part  of  the  world,  are  so  simple  and  natural  that 
no  one  denies  them  ;  but  when  it  is  required  to  state  the 
precise  nature  of  the  morbific  influence  ;  as  we  enter  the 
domain  of  hypothesis,  the  most  various  and  contradic- 
tory opinions  have  been  given,  and  may  be  supported 
with  a  greater  or  less  appearance  of  probability. 

'infectious  diseases  were,  for  a  longtime,  attempted  to 

*  There  is  an  English  translation,  by  W.  Crookes,  London. 


\ 


250 


ON  FERMENTATION. 


be  explained  by  intra-organic  fermentations,  set  up  by 
foreign  bodies  which  everything  induced  us  to  consider 
as  of  an  organized  nature  ;  but  at  a  time  when  the  ideas 
of  fermentation,  properly  so  called,  were  vague  and  ill- 
defined,  it  was  difficult  to  maintain  with  certainty  such 
a  doctrine. 

"  It  has  long  appeared  to  us  that  Pasteur*s  researches 
on  this  subject  have  not  only  succeeded  in  determining 
with  greater  precision  than  before  facts  up  to  that 
time  partly  known,  but  that  they  are  destined  in  the 
future  to  throw  a  bright  light  on  etiology,  and  on  the 
pathological  history  of  contagious  diseases,  whether 
epidemic  or  endemic.  Yet  we  should  not  have  ventured 
to  discuss  here  a  personal  conviction,  which,  though 
shared  by  many  physicians  and  observers,  only  rested 
on  analogy,  were  it  not  that  a  recent  discovery,  due  to 
the  skilful  investigation  of  M.  Davaine,  had  corroborated 
this  opinion  by  a  positive  fact  scientifically  determined, 
a  fact  which  will  certainly  not  remain  unsupported  by 
others. 

"  We  set  out  with  the  hypothesis  that  the  greater  part 
of  infectious  diseases  have  for  their  immediate  cause  the 
penetration  into  the  organism,  and  the  development  of 
the  living  germs  of  ferments,  or  living  ferments  already 
formed,  of  either  animal  or  vegetable  nature,  and  we  will 
make  use  of  the  knowledge  already  acquired  in  order  to 
support  this  opinion  with  a  certain  amount  of  proba- 
bility. We  must  premise,  however,  that  it  will  be 
necessary,  before  we  decide  this  question  definitively, 
to  wait  for  a  direct  and  experimental  proof,  such  as  M. 
Davaine  has  given  with  reference  to  the  "  blood  of  the 
spleen"  or  malignant  boils.  We  think  that  researches  on 


RESEARCHES  OF  M.  PASTEUR. 


251 


fermentation  and  putrefaction  have  been  carried  far 
enough  to  allow  important  trials  to  be  made  in  this 
direction  with  some  hope  of  success.'* 

During  the  ten  years  since  this  page  was  written, 
skilful  experimentalists,  guided  by  the  same  ideas,  among 
whom  I  may  mention  Pasteur  himself  (researches  during 
the  cholera  epidemic),  have  studied  this  subject  with 
great  care,  and  yet  I  must  admit  that  there  has  been  no 
result  from  these  inquiries  ;  the  question  of  the  etiology 
of  infectious  diseases  has  made  no  important  advance ; 
the  observation  made  by  M.  Davaine  remains  without 
any  additional  support. 

Are  we  then  to  conclude  that  these  attractive  predic- 
tions are  erroneous,  and  must  be  rejected,  or  that  these 
observations  are  impossible,  even  with  the  increased 
microscopical  power  which  we  possess  ?  It  is  difficult 
to  decide  one  way  or  the  other,  and  every  question  re- 
quires for  its  decision  positive  facts  ;  negative  results 
can  only  serve  as  a  means  of  checking  our  observed 
facts. 


i 


J 


BOOK  11. 


ALBUMINOID    SUBSTANCES-SOLUBLE   OR   INDIRECT  FER- 
MENTS-ORIGIN OF  FERMENTS. 


CHAPTER  L 

ALBUMINOID  SUBSTANCES,  OR  PROTEIDS. 

Albuminoid  substances,  or  proteids,  play  so  important 
a  part  in  biological  phenomena  in  general,  and  in  the 
nutrition  of  ferments  in  particular,  that  it  would  be 
impossible  for  us  not  to  devote  a  few  pages  to  the 
study  of  these  bodies. 

If  these  substances  represented  chemical  species  well 
defined  and  correctly  classed  among  organic  com- 
pounds— in  other  words,  if  their  constitution  were 
thoroughly  known,  we  should  merely  refer  the  reader  to 
works  on  pure  chemistry,  as  we  have  already  done  in 
the  case  of  fermentable  sugars.  But  unfortunately,  in 
spite  of  many  important  investigations,  it  must  be  said 


254 


ON  FERMENTATION. 


that  the  history  of  proteids  is  still  one  of  the  most 
obscure  subjects  of  organic  chemistry,  one  of  the  most 
urgent  desiderata  of  biological  science. 

So  long  as  the  question  of  the  constitution  of  the 
immediate  principles  of  animal  tissues  has  not  been 
determined,  it  will  be  in  vain  for  physiological  chemistry 
to  investigate  by  the  most  careful  direct  analysis  the 
different  elements  of  an  organ,  or  of  a  liquid,  whether 
in  a  normal  or  pathological  condition  ;  we  shall  always 
be  stopped  by  the  unknown  in  the  interpretation  of 
the  results  which  have  been  obtained.  The  many  and 
various  reactions  which  take  place  in  the  organism 
may  be  regarded  as  true  fermentations,  in  which  the 
fermentable  bodies  are  partly  represented  by  proteids. 
It  is  not  necessary  to  go  into  this  at  great  length  to 
make  it  understood  how  much  these  reactions,  as  observed 
in  the  different  active  phases  of  an  organ,  will  gain 
in  scientific  value,  when  we  are  able  to  formulate  them 
by  means  of  an  equation,  as  we  give  the  formula  of 
alcoholic  fermentation. 

We  shall  study  albuminoid  substances,  by  taking 
this  general  view  of  them,  and  by  bringing  out  more 
prominently,  among  the  results  obtained,  those  which 
throw  some  light  on  the  constitution  and  mode  of 
decomposition  of  these  bodies. 

The  existence  of  a  certain  number  of  albuminoid 
substances,  considered  as  distinct  species,  has  been 
admitted,  being  founded  on  more  or  less  important 
differences  in  their  physical  or  chemical  properties,  or 
in  their  elementary  composition.  This  division  and 
classification  may  be  made  by  two  methods.  If  we 
notice  only  very  decided  differences  of  character  and 


ALBUMINOID  SUBSTANCES,  OR  PROTEIDS.  255 


composition,  we  may  form  out  of  albuminoid  substances 
different  groups  or  families  united  by  common  bonds, 
but  sufficiently  distinct  for  confusion  or  error  to  be 
rendered  impossible. 

Among  these  groups  many  closely  allied  species  may 
be  placed,  which  owe  their  existence  and  individuality 
only  to  certain  subtile  divergencies  of  properties  which 
present  to  the  mind  nothing  very  distinct,  such  as 
the  manner  of  forming  precipitates  with  different 
reagents. 

Without  entering  into  these  details,  we  will  take  as 
types  of  each  group  the  most  important  principle  from 
a  biological  point  of  view.  It  cannot  be  said  that 
substances  to  which  a  special  name  has  thus  been  given, 
as  the  albumin  of  the  egg,  casein,  and  fibrin,  are  imme- 
diate well-defined  principles  ;  for  here  especially  we 
have  no  criterion  by  means  of  which  we  are  able  to 
establish  a  chemical  species.  We  may,  indeed,  suppose, 
with  a  certain  amount  of  probability,  that  they  are 
only  mixtures,  in  variable  proportions,  of  bodies  very 
nearly  allied  and  almost  identical,  the  separation  of 
which  from  each  other  would  be  very  difficult,  if  not 
impossible.  The  natural  fatty  bodies  give  us  in- 
stances of  this  complicated  combination  of  products 
very  similar  in  their  composition  and  character,  and 
which  direct  analysis  has  scarcely  any  power  to 
separate. 

This  hypothesis  has  also  derived  some  support  from 
exact  observations.  Thus,  albumin,  which  was  long 
considered  as  an  immediate  principle,  is,  in  fact,  only 
compounded  of  many  albumins,  having  very  nearly  the 
same  composition,  and  which  can  only  be  distinguished 


256 


ON  FERMENTATION. 


from  each  other  by  their  rotatory  power,  and  by  the 
temperature  at  which  they  coagulate. 

This  question  can  only  be  settled  by  a  careful  study 
of  the  products  of  the  analysis  and  decomposition  of 
albuminoid  substances  ;  as  the  nature  of  fatty  bodies 
has  only  been  understood  by  examining  the  products  of 
their  saponification,  which  are  more  easily  separated 
than  are  the  original  bodies. 

These  views,  which  are  verified  more  and  more,  were 
developed  by  M.  Bouchardat  (Thes  pour  le  concours 
d'agregation,  1872),  and  by  M.  Berthelot,  after  a  com- 
munication made  to  the  "  Societe  Chimique "  by  the 
author  of  this  book.  They  are  very  reasonable,  and 
capable  of  wide  application. 

The  nitrogenous  principles  which  enter  essentially 
into  the  constitution  of  the  organs  and  liquids  of  the 
animal  and  vegetable  economy  are  naturally  divided 
into  various  families. 

The  first  includes  albuminoid  substances,  properly  so 
called  ;  that  is  to  say,  bodies  the  most  nearly  allied,  by 
their  chemical  composition  and  by  the  whole  of  their 
properties,  to  the  albumin  of  egg. 

The  second  is  formed  of  more  remote  products, 
usually  less  rich  in  carbon  and  containing  more  nitro- 
gen. They  enter  into  the  composition  of  less  vital 
tissues,  that  is  to  say,  those  in  which  the  phenomena 
of  nutrition  and  the  changes  are  more  restricted ; 
of  those  which  do  no  work,  that  is,  do  not  develop 
much  force,  such  as  bony,  cartilaginous,  elastic,  fibrous, 
cellular,  horny,  or  epidermic  tissues.  In  this  family  we 
find  horny  tissue,  or  keratrin,  ossein,  cpidcrmose,  elas- 
ticin,  mucin  or  the  gelatines,  and  the  fibroin  of  silk. 


ALBUMINOID  SUBSTANCES,  OR  PROTEIDS.  257 


The  soluble  ferments,  the  peptones,  are  products  of 
the  decomposition  of  albuminoid  substances,  and  should 
be  classed  separately. 

We  find  in  the  organism,  besides  these,  certain  mixed 
principles  placed  between  hydrocarbons  and  proteids, 
true  complex  nitrogenous  glucosides,  decomposing  into 
glucose  and  other  nitrogenous  principles,  such  as  chond- 
rin  and  chitin.* 

Albiuninoid  Substances y  properly  so  called, — Into  this 
family  we  generally  admit  the  following  compounds  : — 

1.  Albumin  of  the  egg,  the  albumin  of  serum  and 
(serine),  vegetable  albumin  ;  these  substances  are  so- 
luble in  water. 

2.  Casein,  vegetable  casein,  paralbumin,  syntonin, 
myosin,  legumin,  amandin,  the  proteids  or  albuminates 
of  the  German  chemists  (this  name  cannot  be  adopted 
in  French,  since  it  gives  an  idea  that  the  body  in 
question  is  a  salt),  paraglobin,  metaglobin  or  fibrino- 
plastic  or  fibrinogenous  substances. 

3.  Blood-fibrin,  coagulated  albumin,  amyloid  matter, 
and  vegetable  fibrin. 

With  regard  to  their  centesimal  composition,  albumi- 
noid substances  are  very  similar  to  each  other ;  yet  the 
analyses  do  not  agree  sufficiently  to  allow  us  to  con- 
clude that  their  composition  is  identical,  or  that  they 
are  isomeric.  We  subjoin  in  a  tabular  form  the  prin- 
cipal analytical  results : — 

• 

*  According  to  my  latest  researches,  albuminoid  substances,  even  those  of 
the  first  family,  contain,  in  small  proportions,  cellulose  amides  as  an  integral 
part  of  their  molecule.  The  distinction  established  between  the  albuminoids 
and  chitin  is  therefore  not  absolute.  The  latter  contains,  it  is  true,  a  greater 
proportion  of  cellulose  amide. 

12 


258 


ON  FERMENTATION. 


Egg  albumin,  not  coagu- 
lated   

Egg  albumin,  not  coagu- 
lated   

Egg  albumin,  purified  ... 

Egg  albumin,  coagulated 

Blood  fibrin   

Casein   

Gluten,     or  vegetable 

fibrin   

Legumin  

Amyloid  matter  

Syntonin  


53*3 

54*3 
52*9 
52*9 
52-8 
53*7 
53*5 
54  *o 

53*1 
53*7 
5o*5 
53*6 
54*1 


7*1 
7*2 
7*2 
7*o 
7*1 
7*1 
7*2 

6-8 
7*2 
6*9 
7*o 
7*2 


15  7 

15-  6 
15*8 

16-  8 

15-  8 
15*8 
15*7 

i5*o 
15*7 
i8-2 
15 'o 

16-  i 


23*6 
22*9 


23  4 
23*4 
23*6 
23"! 


23*4 


Dumas  and  Cahours. 

Scheerer. 

Wurtz. 

Wurtz. 

Dumas  and  Cahours. 
Scheerer. 

Dumas  and  Cahours. 
Scheerer. 

Bou' singault. 
Scheerer, 

Dumas  and  Cahours. 


On  the  one  hand,  it  is  difficult  to  attach  too  great 
importance  to  small  differences,  observed  in  analytical 
results  obtained  with  bodies  so  difficult  to  purify,  amor- 
phous, and  often  containing  mineral  substances  ;  on  the 
other  hand,  nothing  authorizes  us  to  admit  the  complete 
identity  of  their  composition. 

In  fact,  these  bodies  have  certainly  a  very  high 
molecular  weight.  Thus  for  albumin,  by  two  methods, 
(analyses  of  the  potassic  and  platino-hydrocyanic  com- 
binations) nearly  the  same  number,  161 2,  has  been 
arrived  at  for  the  molecular  weight  ;  so  that  if  we  wish 
to  translate  the  results  of  the  elementary  analysis  into  a 
chemical  formula,  as  is  usually  done,  we  are  led  to  very 
high  numbers  in  the  expression  as  in  that  proposed  by 
Lieberkuhn,  C^^  H"'^  W  S  0'''\ 

But  it  is  evident  to  any  one  who  is  in  the  habit  of 
calculating  analyses,  that  a  difference  of  one  atom  of 
carbon,  hydrogen,  or  oxygen,  in  a  formula  with  such 


ALBUMINOID  SUBSTANCES,  OR  PROTEIDS.  259 


high  numbers,  gives  variations  which  are  within  the 
Hmits  of  the  errors  of  analysis.  Elementary  analysis, 
as  we  are  at  present  capable  of  performing  it,  cannot 
solve  the  question  of  isomerism  or  non-isomerism. 

Are  we  farther  advanced  in  that  which  relates  to  the 
constitution  of  these  bodies  ?  At  present,  we  can  foresee 
that,  though  the  question  is  still  in  suspense,  it  will  not 
be  long  before  it  is  determined. 

In  order  to  ascertain  this  constitution,  it  is  necessary 
to  know  exactly  the  whole  of  the  constituent  parts  result- 
ing from  the  destruction  of  proteids,  under  certain  deter- 
minate conditions.  But  in  the  various  reactions  which 
cause  their  splitting  up  and  transformation  into  more 
simple  principles,  we  have  been  able  to  recognize  the 
formation  of  certain  well-defined  compounds  ;  but  these 
bodies  are  more  frequently  accompanied  by  a  relatively 
considerable  number  of  uncrystallizable  bodies,  which 
have  not  yet  been  studied,  and  which  render  any 
attempt  to  determine  an  equation  of  the  reaction 
illusory. 

On  this  account,  we  cannot  form  any  definite  idea  of 
the  manner  in  which  the  72  atoms  of  carbon,  the  112 
atoms  of  hydrogen,  &c.,  of  the  albumin  are  united. 

I  will  give  a  short  summary  of  the  principal  results 
obtained  with  respect  to  these  reactions. 

As  long  since  as  1820,  Braconnot  observed  the  pro- 
duction of  the  sugar  of  gelatin  or  glycocoll,  NO'"^ 
(amido-acetic  acid)  by  boiHng  gelatin  in  sulphuric  acid 
moderately  diluted  ;  by  substituting  muscle  for  gelatin 
he  obtained,  under  the  same  conditions,  the  leucine, 
C«       NO^  (amido^caproic  acid). 

Liebig  afterwards  showed  that  another  crystallizable 


26o 


ON  FERMENTATION. 


product  is  formed  at  the  same  time,  tyrosine,  H"  NO^ 
(oxyphenyl-amido-proprionic  acid). 

Erlenmeyer  and  Schaeffer,  by  extending  these  re- 
searches (the  action  of  boihng  diluted  sulphuric  acid), 
to  most  of  the  albuminoid  substances,  observed  the 
constant  formation  of  leucine  and  tyrosine. 

They  obtained  for  lOO  parts  of  dried  matter — 

Leucine.  Tyrosine. 

Fibrin       .       .       .       .    14     .  .  '8 

Albumin    .       .      .       ,    10     ,  .  I'o 

Syntonin    .       .       .       ,    18     .  .  I'o 

Casein  gives  leucine  and  tyrosine,  and  a  sirupy  re- 
siduum. 

Ritthausen  obtained  by  the  same  kind  of  reactions, 
from  products  formed  by  the  action  of  boiling  dilute 
sulphuric  acid  on  vegetable  nitrogenous  substances,  such 
as  gluten,  two  well  defined  crystallizable  acids. 

Coaglutin  yielded  an  acid  of  the  formula  NO^ 
(glucamic  acid),  homologous  to  aspartic  acid. 

Legumin  gives,  under  the  same  circumstances,  legumic 
acid,  C'H**N'0^* 

According  to  Hlasiwetz  and  Haberman,  the  greater 
part  of  animal  and  vegetable  proteids  are  capable  of  pro- 
ducing these  nitrogenous  acids  (aspartic,  glutamic),  when 
boiled  with  dilute  sulphuric  and  hydrochloric  acids. 

I  have  lately  been  led  to  study,  with  great  care  and 
in  all  its  details,  a  reaction  which  allows  the  albuminoids 
to  be  almost  entirely  resolved  into  crystallizable  princi- 
ples.   My  first  experiments  were  made  in  order  to  ascer- 

»  Ritthausen  says  that  he  has  since  found  that  le^^umic  acid  is  only  a  mix- 
ture of  glutamic  and  aspartic  acids.  I  do  not  think  that  this  opinion  is 
concct. 


ALBUMINOID  SUBSTANCES,  OR  PROTEIDS.     26 1 

tain  whether  a  part  of  the  nitrogen  of  proteinic  com- 
pounds was  not  found  in  the  state  of  urea,  and  whether 
this  class  of  bodies  does  not  represent  complex  ureids. 

After  having  vainly  sought  for  urea  among  the  pro- 
ducts of  the  physiological  decomposition  of  proteids, 
while  ferment  was  kept  without  nourishment,  I  boiled 
albumin,  casein,  &c.,  with  barium  hydrate,  in  an  appa- 
ratus so  arranged  as  to  allow  the  ebullition  to  continue 
during  several  successive  days,  without  any  diminution 
of  the  water.  I  could  easily  accomplish  this,  by  heating 
it  in  a  retort,  attached  to  a  condenser  so  placed  as  to 
allow  the  water  to  return  ;  the  latter  was  connected  with 
two  Wolfe's  bottles,  containing  a  known  volume  of 
normal  sulphuric  acid,  intended  to  retain  the  ammonia 
disengaged. 

Under  these  conditions  at  100°  C.  (212°  F.),  it  was 
found  that  during  the  first  few  hours  there  was  an  abun- 
dant disengagement  of  ammonia,  which  diminished  by 
degrees,  and  at  last  became  imperceptible. 

Nasse  had  previously  observed  this  fact,  and  had 
for  this  reason,  divided  the  nitrogen  of  the  albuminoid 
substances  into  two  portions  ;  one,  the  least  important 
in  quantity  would  be  found,  as  he  stated,  feebly  fixed 
or  combined  {Losegebimdeiier  Sticks  toff). 

The  barium  experiment  gives  us  the  key  to  this 
difference.  In  fact,  at  the  same  time  that  the  ammonia 
is  set  free,  we  see  a  granular  precipitate  formed  in  the 
liquid  which  was  originally  clear  ;  this  precipitate  in- 
creases progressively  to  a  certain  limit,  and  then  ceases 
to  be  formed. 

It  is  almost  entirely  composed  of  barium  carbonate, 
mixed  with  a  small  quantity  of  barium  sulphite  and 


2.62 


ON  FERMENTATION. 


oxalate,  with  silica  derived  from  the  glass  attacked 
by  the  alkaline  liquid,  and  with  alkalino-earthy  phos- 
phates contained  in  the  albuminoid  matter  used  in  the 
experiment. 

By  measuring  the  ammonia  and  barium  carbonate 
formed,  after  boiling  for  120  hours,  there  were  found  in 
100  parts  by  weight  of  dry  albumin — 


By  calculating  the  weight  of  carbon  dioxide  which 
corresponds  to  the  barium  carbonate,  we  find  that  the 
quantities  of  carbon  dioxide  and  ammonia  set  free,  are 
almost  exactly  in  the  proportion  which  urea  would  yield, 
by  resolution  into  ammonium  carbonate  : — 


We  have,  then,  reason  to  admit  the  presence  of  the  same 
grouping  of  elements  as  in  urea  in  albuminous  sub- 
stances. 

Boiling  at  100°  C.  (212°  F.),  even  prolonged  for  eight 
days,  does  not  entirely  destroy  this  grouping  ;  but  albu- 
min is  resolved,  under  the  influence  of  barium  hydrate, 
into  several  more  simple  combinations,  possessing  various 
degrees  of  stability. 

When,  however,  the  temperature  is  raised  to  140°  or 
150°  C.  (284°  or  302°  F.),  the  decomposition  is  complete 
in  a  few  hours  (12  to  24),  and  the  quantities  of  ammonia 
and  carbon  dioxide  remain  the  same,  even  if  we  prolong 
the  operation,  and  raise  the  temperature  of  the  mixture 


Ammonia 
Barium  carbonate 


Gram. 
II'I 


CH*  N2  O  +  K2  O  =  CO2  -f  2  NH3 


ratio  of 


ALBUMINOID  SUBSTANCES,  OR  PROTEIDS.  263 

to  200°  C.  (about  391°  F.).  These  experiments  must 
necessarily  be  made  in  a  close  vessel. 

Under  these  conditions,  we  find  in  100  parts  of  dry 
albumin — 

Ammonia  disengaged     •      •     4*2      •       .  4*5 
Barium  carbonate  .      •       •25        .      •  29 

These  numbers  give  also  the  ratio  of  i'29  between  CO* 
and  NH-l 

Albumin  must  therefore  contain,  in  18  atoms  of 
nitrogen,  about  4  belonging  to  the  urea  group. 

M.  Bechamp  has  noticed,  among  the  products  of 
the  oxidation  of  albumin  by  potassium  permanganate, 
the  presence  of  a  small  quantity  of  urea.  This  fact, 
which  was  denied  by  the  German  chemists,  has  been 
confirmed  by  Ritter's  experiments.  However  this  may 
be,  the  urea  found  existed  in  but  small  quantity  in  the 
products  of  the  reaction  ;  and  it  is  possible  that  its 
production  is  the  result  of  decomposition,  making  that 
which  is  already  formed  sensible  rather  than  the  result 
of  an  oxidation. 

The  barytic  liquid,  separated  from  the  ammonia  and 
from  the  barium  carbonate,  and  freed  from  the  baryta  in 
excess  by  a  current  of  carbon  dioxide,  retains  but  little 
colour  ;  by  concentration  and  crystallization,  followed  by 
treatment  of  the  mother-liquor  with  alcohol,  we  succeed, 
at  last,  in  bringing  it  entirely  into  a  crystalline  form,  that 
is  to  say,  to  the  state  of  a  definite  body.  We  may,  there- 
fore, by  making  a  direct  analysis  of  the  crystals  obtained, 
arrive  at  a  complete  notion  of  the  composition  of  albu- 
minoid substances.  The  reaction  of  the  barium  hydrate, 
at  from  1 50""  to  200°  C.  (302°  to  394°  F.)  gives  us  an  oppor- 


264 


ON  FERMENTATION. 


tunity  of  trying  every  analytical  method  on  the  products 
resulting  from  the  splitting  up  of  albuminoids.  These 
are  not  merely  two  or  three  defined  and  crystallized 
bodies,  which  are  separated  from  a  mass  of  sirupy  matter 
of  considerable  quantity,  which  still  remains  undeter- 
mined ;  on  the  contrary,  we  get  all  the  compounds  that 
are  formed,  and  we  thus  find  ourselves  in  a  position  to 
construct  an  equation  of  the  composition  of  albumin 
and  allied  substances. 

Although  my  researches  on  this  subject  are  not 
entirely  completed,  I  may  already  state,  from  the 
results  obtained, — 

1.  That  barium  hydrate  decomposes  albuminoid  sub- 
stances by  simple  hydratation,  the  experiment  being 
made  in  the  absence  of  oxygen. 

2.  That  the  principal  products  of  this  reaction  are — 
the  elements  of  urea  (ammonia  and  carbon  dioxide,  in 
the  proportion  of  1*29);  traces  of  sulphurous  acid,  of 
sulphuretted  hydrogen,  and  of  oxalic  and  acetic  acids  ; 
tyrosine  H*'  NO'^  (oxyphenyl,  amido-propionic  acid), 
in  very  small  quantity,  two  or  three  per  cent,  of  the 
albumin  at  the  most. 

We  also  find, — 

3.  The  amido  acids  of  the  series  C"  H^^"^'  NO^  cor- 
responding to  the  fatty  acids  H*^"  O*^,  from  amido- 
oenanthylic  acid  C  H'^  NO^  to  amido-propionic  acid; 
leucine  C«  H'^NO^  butalanine  C  H'*  NO' ;  and  amido- 
butyric  acid  C*  IP^  NO'"*  abound  in  this  mixture. 

4.  One  or  two  acids  nearly  allied  to  aspartic  and 
gliUamic  acids  NO*  and  NO* ;  07ie  or  two 
acids  analogous,  and  very  nearly  similar,  to  the  legumic 
acid  found  by  Ritthausen  C«  1 1'*  N'  O^ 


ALBUMINOID  SUBSTANCES,  OR  PROTEIDS.  265 


A  small  quantity  of  a  substance  analogous  to  dextrin, 
which,  by  being  boiled  in  acids,  is  converted  into  a  body 
which  energetically  reduces  Fehling's  liquid  ;  nitric  acid 
changes  it  into  oxalic  acid. 

With  the  exception  of  these  bodies,  nothing  of  im- 
portance is  found  either  as  a  chemical  compound  or  as 
an  amorphous  mass. 

Most  of  the  definite  compounds  found  by  this  reaction 
are  similar  to  those  which  have  already  been  mentioned 
among  the  products  of  the  splitting  up  of  albuminoid 
substances,  under  the  influence  of  acids  ;  but  I  repeat 
that  the  importance  of  the  preceding  reaction  consists 
especially  in  the  demonstration  that  these  compounds 
alone  constitute  the  albuminoid  molecule  ;  and  in  the 
proof  that  the  elements  of  urea  or  carbamide  form  an 
integral  part  of  this  molecule. — (See  the  Memoir  of  the 
author  on  Albuminoid  Substances,  Bulletin  de  la  Soc. 
Chim.  de  Paris,  February  15th,  March  Sth,  and  March 
15th,  1875.) 

The  albumin  is  not  split  up  all  at  once  in  this  manner 
into  comparatively  simple  products  ;  by  stopping  the 
reaction  at  different  periods  of  its  development,  or  by 
using  occasionally  less  elevated  temperatures,  we  meet 
with  intermediate  compounds,  uncrystallizable,  or  crystal- 
lizable  with  difficulty,  the  thorough  study  of  which  will 
be  very  interesting,  in  relation  to  the  phenomena  of 
nutrition  and  of  biological  reactions. 

The  products  which  we  obtain  under  the  influence  of 
the  alkalies  melted  in  their  water  of  crystallization 
(ammonia,  hydrogen,  ammoniacal  compounds,  methyla- 
min,  saniline,  picoline,  petinine,  leucine,  tyrosine,  glyco- 
coll,  carbon-dioxide,  formic,  valerianic,  butyric,  and  oxalic 


266 


ON  FERMENTATION. 


acids)  are  derived  from  a  more  energetic  action,  exerted 
on  the  first  series  of  compounds  formed  by  the  action 
of  baryta.  It  is  the  same  for  the  compounds  which  origi- 
nate under  the  influence  of  oxidizing  agents,  such  as  the 
mixture  of  dilute  sulphuric  acid  and  potassium  bichromate 
(formic,  acetic,  butyric,  valerianic,  caproic,  and  propionic 
acids,  with  their  corresponding  aldehydes,  benzoic  acid 
and  benzoil  hydride,  hydrocyanic  acid  and  butyl  cyanide). 

It  does  not  enter  into  our  programme  to  write  the 
complete  history  of  albuminoid  substances.  (The  reader 
may  consult  with  advantage  on  this  subject  the  Dictionary 
of  Chemistry,  by  M.  Wurtz  ;  FHistoire  Generale  des 
Matieres  Albuminoides,  par  G.  Bouchardat  ;  These 
pour  TAgregation,  Paris,  1872.)  We  shall  merely  give  a 
summary  of  the  theories  proposed,  in  order  to  explain 
the  composition  of  these  complex  bodies. 

All  albuminoid  substances,  heated  for  some  time  with 
alkalies,  dissolve  and  give  up  sulphur  to  the  alkali,  under 
the  form  of  sulphide  and  hyposulphite ;  the  neutraliza- 
tion of  the  liquid  by  an  acid  causes  a  voluminous  floccose, 
white  precipitate,  soluble  in  dilute  lye-water  ;  this  preci- 
pitate has  the  same  composition  whatever  may  be  the 
albuminoid  matter  employed.  Mulder,  relying  on  these 
facts,  considered  this  precipitate  as  the  base  of  albuminoid 
substances,  and  gave  it  the  name  of  protein.  The  Ger- 
man chemist  thought  at  first  that  his  protein  no  longer 
contained  sulphur  ;  later  experiments  have  shown  that  in 
reality  it  still  contains  some,  but  that  this  sulphur  cannot 
be  removed  by  alkalies  under  the  form  of  sulphides. 
According  to  Mulder's  theory,  all  albuminoid  substances 
are  combinations  of  protein  with  variable  quantities  of 
sulphur,  phosphorus,  and  mineral  matter. 


i 


ALBUMINOID  SUBSTANCES,  OR  PROTEIDS.  26/ 

This  opinion,  admitted  very  generally  at  first,  was  by 
degrees  abandoned,  on  account  of  the  great  number  of 
contradictory  observations. 

Liebig  considered  albuminoid  substances  as  having 
the  same  elementary  composition,  ix.y  as  isomeric  com- 
pounds. This  view,  which  admits  of  discussion,  throws 
no  more  light  than  that  of  Mulder  on  what  is  now  called 
the  chemical  constitution  of  these  compounds,  consider- 
ing that  protein  is  almost  as  complex  in  its  character  as 
the  original  products.  Sterry-Hunt  gave  an  explanation 
which  at  first  sight  was  very  seductive ;  he  considered 
albuminoid  substances  as  amides  or  nitrites  of  cellulose, 
dextrin,  gum,  or  sugar. 

The  objection  to  this  simple  idea  is,  that  it  does  not 
agree  with  known  facts  ;  tyrosine,  leucine,  and  aspartic 
acid  are  not  considered  as  derived  from  sugar,  from 
cellulose,  or  from  their  nitrites.  These  latter  bodies 
(nitrites  of  hydrocarbon  substances)  have  been  also  little 
studied,  and  are  little  understood. 

Berthelot,  in  his  treatise  on  elementary  chemistry 
(1872),  considered,  from  the  whole  of  the  facts  then 
known,  albuminoid  substances  to  be  complex  amides, 
formed  by  the  union  of  amido  acids  of  the  series 
CnH^n+i  NO'^(such  asglycocoll  and  leucine),  of  tyrosine, 
with  certain  oxygenated  principles,  some  of  which  belong 
to  the  acetic  series,  and  others  to  the  benzoic  series. 

He  thought  that  the  nature  of  the  amides,  and  of 
the  oxygenized  bodies  which  generated  them,  as  well  as 
their  relative  proportions,  was  the  cause  of  the  differences 
which  exist  between  the  various  albuminoid  bodies ;  and 
that  chitin  and  chondrin  contained,  in  addition,  the 
elements  of  glucose. 


268 


ON  FERMENTATION. 


The  results  which  I  have  obtained  by  means  of  the 
decomposition  by  baryta,  show  that  the  albuminoids  are 
formed  by  the  association,  in  different  proportions,  of  the 
urea  and  arnido-acid  combinations ^  some  belonging  to 
the  series  of  leucine,  H'^^*  NO^  the  others,  which 
are  more  highly  oxygenized,  belonging  to  the  series 
qd.  fj*^n-i  NO*  (aspartic  and  glutamic  acids)  ;  the 
more  complex  acids,  such  as  the  legumic,  may  be 
considered  as  the  products  of  an  incomplete  decom- 
position. Tyrosine  (C^  H"  NO'**)  represents  the  aromatic 
series  ;  it  is  from  this  that  benzoic  and  paroxy-benzoic 
acids  and  bromanil  are  derived,  being  obtained  under 
different  conditions. 

Besides  this,  it  is  easy  to  see  that,  with  the  amido- 
vicids  C'^  H^^"^'  NO^,  it  is  impossible,  even  when  ,  without 
the  urea,  to  arrive  at  the  composition  of  the  albumi- 
noids ;  whatever  combination  of  these  bodies  we  may 
choose,  when  we  deduct  the  elements  of  water  in  suffi- 
cient proportion  to  obtain  the  value  of  the  oxygen  of 
the  albumin,  there  remains  a  very  notable  excess  of 
hydrogen.  The  intervention  of  the  amido-acids  of  the 
aspartic  series  C'^  H*'°~*  NO*,  in  the  grouping  of 
proteids,  is  therefore  indispensable  to  explain  the 
constitution  of  these  bodies. 


CHAPTER  II. 


SOLUBLE  FERMENTS  AND  INDIRECT  FERMENTATION. 

In  the  chemical  phenomena  which  we  have  hitherto 
studied,  we  have  found  the  cause  of  the  reaction  con- 
nected in  such  an  intimate  manner  with  the  presence  of 
a  cellular  organism,  that  all  efforts  made  with  the  view 
of  separating  this  cause  from  this  form  of  life,  even 
for  an  instant,  have  failed,  or  at  least  have  produced  no 
precise  result  definitely  adopted  by  the  scientific  world. 
The  farther  progress  of  science  will,  we  hope,  allow  us 
to  penetrate  more  deeply  into  the  very  essence  of  the 
phenomena. 

If  the  remarkable  and  important  labours  of  Pasteur 
have  taught  us  that  the  transformations  of  sugar  into 
alcohol,  lactic,  and  butyric  acid,  gum  and  mannite, 
those  of  the  albuminoid  substances  into  various  putrid 
principles,  as  well  as  the  conversion  of  alcohol  into 
acetic  acid,  depend  on  the  presence  of  inferior  organisms, 
and  that  the  germs  of  these  organisms  come  from  with- 
out, it  is  no  less  true  that  we  have  as  yet  no  certain 
idea  of  the  mode  of  action  of  organic  ferments.  There 
have  been,  it  is  true,  hypotheses  enough  to  bring  about 
the  solution  of  the  problem.  Liebig,  obliged  to  allow 
more  than  he  had  hitherto  done  to  the  presence  of 
living  organisms,  said  in  his  last  memoir  (Ann.  Chim. 


i 

i 


270  ON  FERMENTATION. 

Phys.  (4),  vol.  23,  p.  6),  that,  in  a  chemical  point  of 
view,  the  only  one  which  he  will  not  give  up,  a  "  vital 
act,"  is  a  phenomenon  of  motion,  and  that  in  this 
sense  M.  Pasteur's  opinion  is  not  contradictory  to  his 
own,  and  is  not  a  refutation  of  it* 

We  have  already  discussed  elsewhere  the  theory  of 
M.  Pasteur,  propounded  as  early  as  1861,  and  lately 
resumed  with  greater  confidence. 

Fermentation  is  a  consequence  of  the  life  of  ferments 
.without  oxygen.  These  simple  organisms  need  oxygen 
so  much,  that  when  they  are  in  a  medium  which  is 
deprived  of  it,  they  take  it  even  from  sugar  and  other 
analogous  bodies.  Fermentation  is  therefore  a  conse- 
quence of  the  disturbance  of  equilibrium  resulting  from 
this  respiration. 

This  manner  of  considering  the  question  is  com- 
pletely tat  variance  with  the  fact  that  the  production  of 
alcohol  at  the  expense  of  sugar,  under  the  influence  of 
a  ferment,  is  not  interfered  with  by  the  presence  of 
oxygen  ;  according  to  Mayer,  it  is  neither  increased  nor 
lessened.  Pasteur  considers  that  it  is  rather  increased 
by  oxygen. 

It  is  certain  that  a  ferment  is  able  to  live  and  develop 
in  a  saccharine,  nitrogenous  and  mineralized  medium, 
without  the  intervention  of  oxygen.    These  same  evolu- 

*  M.  Liebig  aims  here  at  the  following  sentence  of  M.  Pasteur The 
chemical  act  of  fermentation  is  essentially  a  phenomenon  correlative  to  a  vital 
act,  commencing  and  ceasing  with  it.  There  is  never  any  alcoholic  fermenta- 
tion without  there  being,  simultaneously  with  it,  organization,  development^ 
multiplication  of  globules,  or  the  continuous  consecutive  life  of  globules  already 
formed."  According  to  the  German  chemist,  fermentation  is  a  movement 
communicated  by  instable  bodies  in  process  of  chemical  transformation;  it 
signifies  but  little  whether  or  no  these  transformations  take  place  in  a  li\ing 
organism. 


SOLUBLE  FERMENTS. 


271 


tions  are  more  active  under  the  influence  of  air.  The 
only  inference  that  we  can  draw  from  these  well- 
estabHshed  facts  is,  that  up  to  a  certain  point,  the 
sugar,  which  the  ferment  always  resolves  into  alcohol 
and  carbon  dioxide,  as  long  as  it  can  meet  with  it,  and 
unless  it  is  exhausted,  is  able  to  supply  to  the  ferment 
the  living  forces  required  for  its  development.  But  we 
know  not  whether  alcoholic  fermentation  is  a  conse- 
quence of  this  borrowing  of  living  force,  or  whether  it 
is  a  previous  action. 

The  chemical  cause  of  alcoholic  fermentation  set  up 
by  yeast  has  been  sought  for  from  another  point  of 
view,  which  must  now  be  the  object  of  our  studies. 

We  know  that  cane-sugar,  before  it  undergoes  alco- 
holic fermentation,  is  hydrated,  and  thus  split  up,  as 
often  occurs  under  the  influence  of  acids,  into  two 
opposite  glucoses  ;  the  ordinary  glucose,  or  grape-sugar, 
which  causes  the  plane  of  polarization  to  turn  to  the 
right  and  laevulose,  or  uncrystallizable  sugar.  This 
change  was  at  first  attributed  to  the  acidity  of  the  fer- 
ment. M.  Berthelot,  and  after  him  M.  Bechamp, 
showed  that  the  active  agent  is  a  soluble,  neutral,  nitro- 
genous principle,  excreted  by  the  ferment,  which  is 
found  in  greater  or  less  abundance  in  the  filtered  water 
in  which  yeast  has  been  washed  (the  zymase  of  Be- 
champ, the  alterative  ferment  {ferment  inversive)  of 
Berthelot). 

This  soluble  principle,  to  which  we  can  attribute  no 
organization,  but  which  is  directly  derived  from  a  living 
organism,  possesses  the  remarkable  power  of  altering 
cane-sugar  in  a  few  moments. 

When  this  fact  was  established  concerning^  the  altera- 


272 


ON  FERMENTATION. 


tive,  soluble,  inorganic  ferment,  analogous  compounds 
had  been  long  known,  also  soluble,  nitrogenous,  and  inor- 
ganic, which  were  especially  characterized  by  specific 
acts  which  they  could  exercise  chemically  on  various 
principles. 

It  was  thus  that  M.  Payen  and  M.  Persoz  obtained 
from  barley  that  had  germinated,  and  had  been  after- 
wards ground,  by  treating  it  with  water,  a  soluble 
substance  capable  of  turning  starch  into  sugar.  The 
presence  of  emulsin  had  also  been  ascertained  in 
almonds,  and  this  transforms  amygdalin  into  essence 
of  bitter  almonds. 

That  which  especially  distinguishes  these  chemical 
reactions,  set  up  by  these  various  soluble,  inorganic 
principles,  is  the  greatness  of  the  effect  compared  with 
the  trifling  quantity  of  the  active  agent.  This  same 
character  is  found  also  in  direct  fermentation,  due  to 
the  immediate  intervention  of  living  organisms. 

We  will  give  the  name  of  indirect  fermentation  to  the 
reactions  of  which  we  have  just  spoken,  and  whose  cause 
is  derived  from  an  organism,  but  can  act  without  it. 

It  was  natural  to  seek  for  the  cause  of  direct  fer- 
mentation in  the  intervention  of  active  products, 
whether  soluble  or  not,  elaborated  by  organic  ferments. 
Thus  two  orders  of  phenomena,  which  are  certainly 
not  without  relation  to  each  other,  were  connected 
together. 

This  theory,  which  would  cause  the  direct  or  true 
fermentation  of  M.  Pasteur  to  be  confounded,  as  to 
its  essence,  with  indirect  fermentation,  or  fermenta- 
tion caused  by  inorganic  soluble  ferments,  has  not 
found  sufficient  support  from  characteristic  and  well- 


SOLUBLE  FERMENTS. 


observed  facts,  to  be  established  as  a  truth  that  has 
been  demonstrated  ;  but  it  has  not  yet  been  absolutely- 
contradicted.  It  is,  however,  easily  to  be  seen  that 
such  an  explanation  becomes  useless.  If  the  decom- 
posing force  is  derived  from  the  cell,  what  would  be 
the  utility  of  this  intermediate  soluble  or  insoluble 
solid  substance  which  transmits  the  force  ? 

General  Characters  of  Soluble  Ferments, — Soluble  fer- 
ments are  all  derived  directly  from  living  organisms  in 
the  midst  of  which  they  originate.  Up  to  the  present 
time,  the  specific  characters,  of  which  we  shall  presently 
speak,  have  not  been  communicated  to  any  artificial 
organic  substance.  We  are,  therefore,  compelled  to 
believe  that  this  specific  character  is  a  consequence  of 
the  origin  of  soluble  ferm.ents.  Their  composition  resem- 
bles that  of  albuminoid  substances  ;  in  fact,  they  con- 
tain carbon,  nitrogen,  hydrogen,  and  oxygen.  But  the 
analogy  will  go  no  farther.  When  we  have  eliminated, 
by  proper  processes  which  we  are  about  to  describe,  the 
albuminoid  substances  which  always  accompany  soluble 
ferments  in  their  first  solutions,  we  find  that  the  product, 
though  it  preserves  all  its  chemical  activity,  no  longer 
manifests  the  general  reactions  of  albuminoid  sub- 
stances. It  no  longer  yields  a  precipitate  with  tannin 
and  corrosive  sublimate ;  iodine  and  nitric  acid  no 
longer  colour  it. 

Elementary  analyses  have  also  revealed  sensible  dif- 
ferences ;  but  as  there  was  no  proof  that  the  products 
analyzed  were  pure,  we  can  draw  no  conclusion  from 
this  concerning  the  chemical  nature  of  soluble  ferments. 
It  is  probable  that  they  are  derived  from  the  physio- 
logical splitting  up  of  proteids. 


274 


ON  FERMENTATION. 


The  sohible  indirect  ferments,  or  zymases^  present 
themselves,  when  in  a  dry  state,  as  amorphous,  colour- 
less, pulverulent  matter ;  they  are  usually  precipitated 
from  their  aqueous  solutions  by  alcohol,  corrosive  subli- 
mate, neutral  or  basic  lead-acetate.  These  precipitates, 
when  decomposed  by  sulphuretted  hydrogen,  restore  to 
the  water  the  soluble  matter  unchanged,  and  preserving 
its  specific  properties  ;  but  it  is  still,  in  this  case,  accom- 
panied by  a  greater  or  less  quantity  of  albuminoid  matter. 

If  the  sublimate  precipitates  them  from  liquids  ex- 
tracted from  the  organism,  it  is  rather  by  mechanical 
action  than  by  the  fact  of  a  chemical  combination  ;  for 
we  have  just  seen  that  soluble  ferments,  when  deprived 
of  albuminous  principles,  are  not  precipitated  by  the 
sublimate. 

By  taking  advantage  of  the  facility  with  which  the 
zymases  are  mechanically  carried  down  by  solid  deposits 
in  process  of  formation  in  the  midst  of  the  liquid  which 
contains  them,  certain  chemists  have  discovered  means 
of  purifying  them.  Thus  Conheim  separates  pure 
ptyalin  (salivary  diastase)  by  acidulating  the  saliva 
strongly  with  tribasic  phosphoric  acid  ;  the  phosphoric 
acid  is  subsequently  neutralized  by  lime-water,  until  an 
alkaline  reaction  takes  place.  The  precipitate  of  tri- 
calcic  phosphate  carries  down  with  it,  as  it  is  formed,  the 
ptyalin  and  albuminoid  matter ;  the  filtered  liquid  has 
no  action  on  starch.  The  deposit,  washed  with  water, 
yields  the  ptyalin  to  the  solvent,  and  retains  the  pro- 
teinic matter ;  it  is  then  only  necessary  to  precipitate  it 
by  alcohol,  and  we  obtain  a  white,  light,  flaky  deposit, 
which,  dried  in  vacuo,  presents  itself  in  the  form  of  an 
almost  colourless  powder. 


SOLUBLE  fermp:nts. 


275 


Pure  pepsin  is  extracted  in  a  similar  manner  from 
natural  or  artificial  gastric  juice  (obtained  by  exposing 
the  mucous  membrane  of  the  stomach,  separated  from 
the  muscular  membrane,  and  cut  up,  to  a  temperature 
of  35°  C.  (95°  F.),  with  water  containing  5  per  cent,  of 
phosphoric  acid.  This  juice,  which  contains  phosphoric 
acid,  is  precipitated  by  lime-water.  The  tricalcic  phos- 
phate is  transformed,  by  the  addition  of  a  proper  quan- 
tity of  phosphoric  acid,  into  insoluble  and  crystallized 
bicalcic  phosphate,  which  it  is  only  necessary  to  wash  in 
order  to  remove  the  pepsin  adhering  to  it.  We  may  also 
dissolve  the  precipitate  of  calcium  phosphate  in  dilute 
hydrochloric  acid  ;  then  pour  into  the  liquid  a  solution 
of  cholesterin  in  a  mixture  of  four  parts  of  alcohol  and 
one  of  ether ;  shake  up  with  the  liquid  the  cholesterin 
which  is  separated,  and  collect  it  on  a  filter  ;  wash  it 
with  water  acidulated  with  acetic  acid,  then  with  pure 
water.  The  damp  cholesterin,  to  which  the  pepsin 
adheres,  is  treated  with  pure  ether,  which  dissolves  it, 
while  there  remains  in  the  lower  part  of  the  vessel  a 
solution  of  pure  pepsin  in  water  (Briicke).  This  pepsin 
yields  no  precipitate  except  to  platinum  bichloride,  and 
neutral  and  basic  acetates  of  lead.  Nitric  acid,  tannin, 
and  corrosive  sublimate  are  without  effect.  It  only  yields 
a  very  slight  colour  with  nitric  acid  and  ammonia. 

Danilewsky  makes  use  of  collodion  to  precipitate 
soluble  ferments.  The  precipitate,  having  been  well 
washed,  is  treated,  after  being  dried,  with  a  mixture  of 
ether,  alcohol,  and  water,  which  dissolves  the  nitrated 
cellulose,  leaving  in  solution  the  active  principle,  free 
from  albuminoid  matter. 

Von  Wittich  (Arch.  f.  d.  ges.  Physio.,  vol.  3,  p.  339), 


2/6 


ON  FERMENTATION. 


proposes  the  following  method,  applicable  to  the  extrac- 
tion of  soluble  ferments  in  general.  The  vegetable  or 
animal  organ  which  contains  them  is  rapidly  cut  in 
pieces,  cleared  from  blood,  if  necessary,  by  washing  in 
water,  and  then  left  under  alcohol  for  twenty-four  hours  ; 
afterwards  dried  in  the  air,  pulverized  and  sifted. 

The  powder  is  diffused  in  glycerin  ;  and  this  glycerin 
solution  is  precipitated  by  alcohol.  By  repeating  this 
operation  several  times  (solution  in  glycerin  and  precipi- 
tation by  alcohol)  an  active  powder,  free  from  albuminoid 
matter,  is  obtained. 

We  have  but  little  information  respecting  indirect 
ferments,  regarded  in  a  chemical  point  of  view.  We 
cannot  say  whether  or  no  they  have  the  same  composi- 
tion, and  only  differ  by  their  specific  activity.  That 
which  gives  considerable  importance  to  these  products 
is  the  transforming  power  which  they  exercise  over  a 
great  number  of  organic  compounds,  and  the  important 
part  which  they  play  in  many  physiological  reactions. 
The  activity  of  an  indirect  ferment  depends  on  the 
temperature,  like  that  of  organic  ferments.  In  general 
terms,  we  may  say  that  it  increases  with  the  temperature 
up  to  a  certain  limit,  beyond  which  it  undergoes  a  rapid 
depression,  till  it  ceases  altogether.  This  limit  varies 
with  the  nature  of  the  ferment ;  it  is  always  under 
ioo°  C.  (212°  F.),  and  is  found  to  be  higher  than  that 
of  organic  ferments. 

The  action  of  chemical  agents  on  soluble  ferments 
is  also  not  quite  to  be  compared  with  that  exerted  on 
organic  ferments. 

Thus  M.  P.  Bert  has  observed  that  compressed 
oxygen  destroys  the  latter  ferments  after  a  longer  or 


SOLUBLE  FERMENTS. 


277 


shorter  interval,  while  soluble  ferments  are  not  modi- 
fied in  their  activity.  This  interesting  experiment 
establishes  a  very  distinct  line  of  demarcation  between 
the  two  kinds  of  ferments,  and  may  serve  to  classify 
them,  whenever  microscopical  indications  leave  any 
doubt  as  to  their  character. 

M.  Bouchardat  (Ann.  Chim.  Phys.  (3),  vol.  14,  p.  61), 
who  has  carefully  studied  the  influence  of  chemical 
compounds  on  diastase,  has  observed  that  certain  sub- 
stances, which  are  antagonistic  to  alcoholic  fermenta- 
tion, have  no  influence  on  the  effects  of  diastase ;  such 
as  prussic  acid,  the  mercurial  salts,  alcohol,  ether,  chloro- 
form, and  certain  essences  (cloves,  turpentine,  lemon, 
mustard,  &c.)  Citric  and  tartaric  acid,  which  only 
slightly  interfere  with  alcoholic  fermentation,  completely 
destroy  the  activity  of  diastase. 

M.  Dumas  (Comp.  Rend.,  vol.  75,  p.  295),  made 
special  experiments  on  the  action  of  borax  on  this 
class  of  ferments.  He  found  that  a  solution  of  borax 
coagulates  beer-yeast ;  the  supernatant  liquid  has  lost 
the  property  of  altering  cane-sugar ;  it  also  neutralizes 
the  action  of  the  water  of  yeast  on  saccharose.  If  we 
place  sweetened  Avater  and  the  water  of  yeast  in  one 
tube,  and  sweetened  water  with  yeast-water  and  a 
solution  of  borax  in  another,  the  first  will  soon  show 
signs  of  alteration,  while  the  second  will  manifest  none. 
Analogous  eff*ects  are  observed  with  synaptase  or  emulsin, 
diastase,  and  myrosin.  All  these  ferments  cease  to  act 
on  amygdalin,  starches,  and  myronic  acid,  from  the 
moment  that  they  are  placed  in  contact  with  a  solution 
of  borax.  This  salt  appears,  then,  to  have  a  specific 
action  in  destroying  the  activity  of  all  soluble  ferments. 


278 


ON  FERMENTATION. 


We  have  seen,  on  the  contrary,  that  yeast,  placed  in 
contact  for  three  days  with  a  saturated  solution  of 
borax,  is  able  still  to  set  up  alcoholic  fermentation. 
Borax  may  then,  like  compressed  oxygen,  serve  as  a 
differentiating  character  of  soluble  and  organic  ferments. 

BodieSy  or  Chemical  Compounds,  on  which  Soluble  Per- 
mcjits  are  able  to  Act,  and  the  Kind  of  Reactions  which 
they  Excite, — Organic  ferments  exert  their  activity  on 
a  great  number  of  organic  compounds  belonging  to 
different  groups. 

The  glucoses,  acids  rich  in  oxygen,  such  as  malic, 
tartaric,  citric  or  lactic  acids,  albuminoid  substances, 
urea,  and  alcohol,  can  be  affected  by  organic  living  fer- 
ments. The  reactions  which  these  ferments  cause  them 
to  undergo  are  often  very  complex,  and  cannot  be  for- 
mulated by  simple  equations,  proVided  that  we  wish  to 
represent  correctly  all  the  terms ;  in  fact,  in  almost 
every  case,  we  have  not  been  able  to  reproduce  arti- 
ficially the  complex  conditions  of  organic  matter  under 
the  influence  of  organic  ferments. 

Indirect  or  soluble  ferments  are  able  to  act  also  on 
various  classes  of  organic  compounds,  but  the  mode 
of  action  is  generally  the  same.  There  is  a  more  or 
less  simple  splitting  up  accompanied  by  a  hydrata- 
tion.  The  nature  of  this  splitting  up  is  always  con- 
formed to  the  peculiar  constitution  of  the  compound, 
and  may  be  explained,  in  most  cases,  by  chemical  pro- 
cesses in  which  the  direct  or  indirect  intervention  of  a 
living  organism  cannot  be  brought  in. 

Thus  starch  is  resolved  by  hydratation  into  glucose 
and  dextrin,  and  this  in  its  turn  is  converted  into 
maltose,  as  well  under  the  influence  of  diastase  as 


SOLUBLE  fermj:nts. 


279 


by  being  boiled  with  a  dilute  acid  (sulphuric  acid). 
The  alterative  ferment  hydrates  a  molecule  of  saccha- 
rose, and  converts  it  into  two  molecules  of  glucose : 
dilute  acids  behave  in  the  same  manner. 

Certain  special  soluble  ferments,  such  as  synaptase 
or  emulsin  contained  in  sweet  and  bitter  almonds,  and 
the  myrosin  of  white  or  black  mustard,  act  on  natural 
glucosides  ;  that  is  to  say,  on  those  special  compounds 
that  are  met  with  in  plants  in  such  great  abundance, 
and  which  we  may  consider  as  ethers  composed  of 
glucoses,  or  of  polyatomic  alcohols.  The  result  is  a 
splitting  up,  by  hydratation,  into  glucose  and  another 
principle.  Chemical  means,  such  as  boiling  with  an 
acid  or  an  alkali,  lead  to  the  same  result. 

The  fatty  bodies  (ethers  compounded  with  glycerin), 
as  is  well  known  by  Chevreul's  beautiful  experiments, 
fix  the  elements  of  water  when  they  are  heated  with 
boiling  alkalies,  or  with  acids,  and  yield  glycerin  and  a 
fatty  acid,  the  sum  of  the  weights  of  which  is  equal  to 
the  weights  of  fatty  matter  used,  plus  the  water  fixed 
by  the  reaction. 

But  we  know  by  the  works  of  M.  C.  Bernard,  that 
the  pancreatic  gland  secretes  a  soluble  nitrogenous 
substance,  capable,  like  boiling  alkalies,  of  saponifying 
greasy  substances. 

It  is  very  probable  that  the  digestion  of  albuminous 
matters,  and  their  conversion  into  peptones,  under  the 
influence  of  the  gastric  or  pancreatic  juice,  or  rather 
under  that  of  soluble  ferments  contained  in  the  secre- 
tions, are  only  the  results  of  a  hydratation  and  a 
splitting  up,  the  conditions  of  which  we  are  able  to 
realize  without  the  agency  of  life. 


280 


ON  FERMENTATION. 


We  may  almost  foresee  that,  in  general,  every  pheno- 
menon of  the  splitting  up  of  an  organic  compound 
into  its  proximate  constituent  parts,  requiring  only  a 
simple  fixation  of  water,  will  find  in  the  organism 
itself  its  soluble  ferment,  that  is  to  say,  the  agent 
capable  of  producing  this  decomposition.  It  is  thus 
that  ethers  composed  of  mon-atomic  or  polyatomic 
alcohols,  including  the  glucosides  and  fatty  bodies, 
certain  complex  acids,  such  as  the  hippuric,  glyco- 
cholic,  taurocholic,  &c.,  are  resolved  into  two  or  more 
simpler  molecules. 

Can  the  same  soluble  ferment  act  upon  different 
chemical  compounds,  decomposing  them  in  the  manner 
indicated  by  the  peculiar  constitution  of  each  ?  There 
is  not  the  shadow  of  a  doubt  as  to  the  positive  answer 
to  this  question.  Thus  we  see  emulsin  or  synaptase, 
contained  in  almonds,  cause  the  decomposition  of  a 
great  number  bf  crystallizable  and  definite  principles 
of  the  vegetable  organism,  such  as  amygdalin,  salicin, 
arbutin,  helicin,  phlorizin,  esculin,  daphnin. 

It  is  true  that  these  various  bodies  belong  to  the 
same  family,  and  have  the  same  functions ;  they  are 
all  glucosides,  and  we  must  not  be  more  surprised  to 
see  the  same  force  transform  them  than  to  observe  the 
saponification  of  neutral  fatty  bodies  under  the  same 
influences. 

In  certain  cases,  we  see  the  same  organic  liquid,  the 
same  secretion,  such  as  the  pancreatic  juice,  exert  its 
active  power  upon  very  different  principles,  which  do  not 
present,  as  in  the  previous  example,  an  analogous  con- 
stitution. The  pancreatic  juice  transforms,  modifies, 
and  digests  albuminoid  substances,  turns  starch  into 


SOLUBLE  FERMENTS. 


28l 


sugar,  and  saponifies  fatty  substances.  We  may  well 
ask  if  this  multiple  specific  power  belongs  to  one  and 
the  same  principle,  or  whether  it  indicates  the  presence 
of  several  distinct  soluble  ferments  in  the  pancreatic 
juice. 

The  experiments  of  Cohnheim  and  Danilewsky  tend 
to  prove  that  the  various  physiological  properties  of  the 
pancreatic  juice  are  due  to  special  principles.  These 
chemists  have,  indeed,  been  able  to  separate  by  precipi- 
tation, by  means  of  calcium  phosphate  or  collodion,  a 
nitrogenous  substance,  which  is  not  albuminoid,  and 
which  shows  all  the  characters  of  salivary  diastase,  and 
which  rapidly  turns  starch  into  sugar,  without  digesting 
the  proteids. 

Thus,  when  we  pour  the  ethereal  alcoholic  solution  of 
collodion  into  the  extract  of  the  gland,  obtained  by 
pounding  it  with  magnesium  carbonate  and  water,  and 
afterwards  filtering  it,  the  ferment  of  albuminoid  sub- 
stances is  precipitated  with  the  collodion,  while  the 
amylaceous  ferment  remains  in  the  liquor,  and  may 
be  obtained  by  evaporation.  Calcium  phosphate  preci- 
pitated from  an  aqueous  solution  of  the  pancreatic 
gland  (by  phosphoric  acid  and  lime-water),  carries  down 
with  it  the  diastasic  ferment ;  while,  on  the  contrary, 
the  albuminoid  ferment  remains  in  solution.  If,  in  the 
experiment  made  with  collodion,  the  precipitate  be  redis- 
solved,  washed  in  water,  and  dried  in  aqueous  ether,  we 
obtain  two  layers  ;  the  lower  and  watery  one  contains 
the  albuminoid  ferment  in  solution.  As  to  the  ferment 
which  saponifies  the  fatty  matters,  it  has  not  yet  been 
obtained  by  itself,  and  independently  of  the  two  others. 

Lhnits  of  the  Activity  of  a  Sohtble  Ferment. —  Certain 
13 


282 


ON  FERMENTATION. 


experiments  tend  to  prove  that  soluble  ferments  have 
not  an  unlimited  activity.  It  is  known  that,  with  a 
determinate  weight  of  one  of  these  bodies,  we  can 
modify  an  incomparably  greater  quantity  of  fermentable 
matter  ;  but  does  the  ferment  always  lose  its  power  after 
a  time,  and  become  exhausted  ?  The  Hmit  of  activity 
has  been  determined  for  only  a  small  number  of  these 
ferments,  and  the  results  published  warrant  but  very 
undecisive  conclusions.  Most  of  the  experiments  of  this 
kind  have  been  made  with  impure  ferments,  still  con- 
taining albuminoid  substances. 

Thus,  according  to  Payen  and  Persoz,  one  part  only 
of  diastase  is  sufficient  to  liquefy  and  turn  to  sugar  2,000 
parts  of  starch  ;  but,  on  the  one  hand,  the  diastase  pre- 
pared by  alcoholic  precipitation,  according  to  the  process 
employed  by  these  observers,  is  a  complex  mixture,  into 
which  the  really  active  substance  enters,  perhaps,  only  in 
a  very  small  proportion.  The  ratio  of  I  to  2,000  would 
thus  become  much  smaller.  On  the  other  hand,  it  has 
been  ascertained  that  the  presence  of  a  certain  quantity 
of  glucose  interferes  with  the  transformation  of  dextrin  ; 
and  in  order  to  allow  the  action  to  resume  its  course, 
it  is  only  necessary  to  dilute  the  liquid  with  much  water, 
or  to  remove  the  glucose  as  it  is  formed,  by  causing  it 
to  undergo  an  alcoholic  fermentation  by  means  of 
yeast. 

/  According  to  M.  Berthelot,  the  alterative  ferment  of 
yeast  is  able  to  alter  from  50  to  100  times  its  weight  of 
sugar.  We  ought  to  make  the  same  remark  about  these 
numbers  as  v/e  did  respecting  diastase.  Nothing,  there- 
fore, proves  decidedly  that  soluble  ferments  lose  their 
specific  power  as  they  exercise  it.    The  opposite  opinion 


SOLUBLE  FERMENTS. 


283 


finds  some  support  in  the  infinitely  small  quantities  of 
the  ferments  necessary  to  produce  a  very  important 
and  considerable  effect.  It  is  also  evident  that  these 
decompositions  are  effected  rather  with  disengagement 
than  absorption  of  heat,  since  the  same  decomposition 
by  hydratation  takes  place  by  the  action  of  sulphuric 
acid  in  small  quantities,  and  the  same  amount  of  acid 
will  act  indefinitely.  They  do  not,  therefore,  require  the 
employment  and  the  consumption  of  living  forces,  and 
the  principle  of  the  conservation  of  energy  is  not  op- 
posed to  the  idea  of  an  indefinitely  prolonged  action. 

This  manner  of  considering  the  question  does  not 
exclude  the  possible,  and  even  probable  fact,  shown  in 
certain  cases,  of  progressive  weakening  of  the  ferment, 
after  which  it  will  have  lost  its  specific  power :  a  similar 
chemical  decay  may  accompany  the  manifestation  of 
the  specific  power,  or  be  produced  without  it,  in  an 
independent  manner,  and  without  being  a  consequence 
of  it. 

Particular  Studies  of  Soluble  Ferments  or  Zymases, — 
After  these  general  considerations,  we  will  notice  what 
speciality  is  presented  by  each  indirect  ferment.  In  this 
particular  study,  we  will  not  return  to  the  physical 
characters,  the  chemical  composition  and  properties  of 
zymases,  nor  to  the  method  of  preparing  them  in  greater 
or  less  purity.  These  different  questions  have  been  fully 
treated  in  the  general  study  of  these  bodies,  and  we 
should  expose  ourselves  to  needless  repetitions  by  bring- 
ing forward  again  for  each  ferment  that  which  applies 
to  the  whole  class. 

The  points  which  will  claim  our  attention  are  the 
chemical  reaction,  the  different  sources  of  the  ferment 


2S4 


ON  FERMENTATION. 


which  excite  it,  and  the  special  conditions  which 
favour  it. 

I.  Diastases. — We  shall  give  the  general  name  of 
diastases  to  ferments  capable  of  turning  starch  into 
sugar. 

If  we  only  regard  the  original  and  the  final  terms  of 
the  reaction,  we  arrive  at  this  conclusion,  that  starch, 
under  the  influence  of  diastase,  combines  with  itself  the 
elements  of  water,  and  is  converted  into  glucose. 

C«  H^o  o^  +  H2  O  =  C°  o« 

Amylaceous  matter.    Water.  Glucose. 

Yet,  according  to  the  researches  of  Musculus  (Ann. 
Phys.  Chim.  (3),  vol.  55,  p.  203  ;  Bulletin  de  la  Soc.  Chim., 
Paris,  vol.  23,  p.  31,  1874)  ;  those  of  Schwarzer  (Bull.  Soc. 
Chim.,  vol.  14,  p.  400)  ;  those  of  Schultz  and  Maerker 
(Bull.  Soc.  Chim.,  vol.  19,  p.  17),  and  those  of  Payen,  the 
change  does  not  take  place  so  simply.  By  previously 
warming  the  starch  with  water  at  70"^  C.  (158°  F.),  before 
adding  the  diastase,  the  reaction  by  means  of  iodine 
disappears  at  the  moment  that  the  saccharification  has 
affected  a  quarter  of  the  liquid.*  But  the  action  of 
diastase  does  not  stop  there  ;  if  we  add  a  larger  quantity, 
it  continues  till  it  has  reached  the  half  (5 1  per  cent, 
according  to  Payen  ;  from  51  to  5 17  per  cent,  according 
to  Schultz  and  Maerker)  ;  that  is  to  say,  that  the  starch 
is  converted  into  a  mixture  of  glucose  and  dextrin  in 
equal  equivalents.  From  this  moment  the  saccharifi- 
cation stops,  notwithstanding  the  addition  of  greater 
quantities  of  diastase. 

With  soluble  starch  the  action  is  the  same.  The 


^  Not  the  third,  as  Musculus  had  previously  announced. 


SOLUBLE  FERMENTS. 


285 


reaction  by  iodine  disappears  as  soon  as  a  quantity  of 
glucose,  equivalent  to  a  quarter  of  the  starch,  has  been 
formed  ;  but  with  an  excess  of  diastase,  glucose  is  rapidly 
and  easily  formed  at  the  expense  of  half  the  starch. 

According  to  these  results,  and  without  taking  into 
consideration  the  disappearance  of  the  peculiar  colour 
produced  by  iodine,  when  the  quantity  of  sugar  is  equal 
to  one-quarter,  a  fact  which  must  be  interpreted  by  fresh 
experiments,  we  may  suppose  that  the  amylaceous 
molecule  is  resolved  by  hydratation  into  two  molecules, 
one  of  glucose,  the  other  of  dextrin. 

2  (C^  H'o  O^)  +  H2  O  =  C6  H^o  05  +  a  O^ 
Starch.  V^ater.  Dextrin.  Glucose. 

M.  Payen  has  shown  long  since,  that  by  the  intervention 
of  yeast  we  produce  the  entire  transformation  of  starch 
into  glucose,  and  finally  into  alcohol  and  carbon  dioxide. 

This  experiment  would  prove  that  the  presence  of 
glucose  is  antagonistic  to  the  action  of  diastase  upon 
dextrin. 

Thus  starch,  soluble  starch,  dextrin,  and,  according  to 
the  researches  made  by  CI.  Bernard,  the  glycogen  matter 
of  animal  tissues  can  be  changed  into  sugar  under  the 
influence  of  diastase.* 

*  A  diastase  of  considerable  activity  may  be  obtained  by  a  modification  of 
the  process  of  MM.  Payen  and  Persoz.  One  part  of  germinated  barley  in 
powder  is  added  to  two  parts  of  water.  After  an  hour's  maceration,  the  liquid 
is  pressed  out,  and  an  equal  volume  of  alcohol  at  80°  C.  (176°  F.)  is  added  to 
it.  It  is  then  filtered,  and  the  first  rather  voluminous  precipitate  is  rejected. 
An  equal  volume  of  alcohol  is  again  added  to  the  filtered  liquid.  A  very  slight 
precipitate  is  then  formed,  which  is  collected  on  a  filter;  this  filter  is  dried 
with  the  precipitate  at  a  gentle  heat.  A  diastase  paper,  which  is  very  active,  is 
obtained  in  this  manner,  and  may  be  indefinitely  kept  without  change.  (Mus- 
culus.) 


286 


ON  FERMENTATION. 


Diastasic  reactions  play  an  important  part  in  the 
animal  organism,  and  especially  in  the  digestion  of 
starchy  food.  Thus  we  find  diastasic  ferment  in  the 
animal  economy  as  well  as  in  the  vegetable  tissues.  At 
its  first  entrance  into  the  digestive  tube,  the  food  is  mixed 
with  the  saliva,  a  liquid  which  contains,  as  shown  by 
M.  Mialhe  in  1845,  ptyalin^  a  fermenting  principle  ana- 
logous, or  rather  identical,  with  the  diastase  of  germinated 
barley.  MM.  Bouchardat  and  Sandras  proved  nearly 
at  the  same  time  the  existence  of  an  analogous  agent  in 
the  pancreatic  juice. 

The  pancreatic  juice  acts  infinitely  more  energetically 
than  saliva  in  the  saccharification  of  starchy  substances. 
The  action  of  an  infusion  of  pancreatic  juice  upon  starch 
is  excessively  rapid — indeed,  we  may  say,  instantaneous  ; 
it  is,  in  fact,  in  the  small  intestine  that  the  principal  diges- 
tion of  amylaceous  substances  is  effected. 

This  same  ferment  is  met  with  in  other  parts  of  the 
organism  ;  everywhere  where  starch,  whether  animal  or 
vegetable,  must  be  rendered  soluble.  Thus,  in  the  liver 
there  exists  a  kind  of  animal  starch,  glycogen,  which  is 
turned  into  sugar  by  contact  with  blood,  and  a 
ferment,  in  order  to  be  conveyed  in  this  form  into  the 
torrent  of  the  circulation.  At  the  period  of  animal 
life  when  this  change  is  to  be  accomplished,  the  ferment 
appears,  and  the  accumulated  starch  is  destroyed.  M.Cl. 
Bernard  (Digestion  comparee  chez  les  Animaux  et  les 
Vegetaux,  Revue  Scientifique,  1873,  p.  515)  draws  a 
very  striking  parallel  between  the  chemical  phenomena 
of  nutrition  in  animals  and  vegetables. 

"  The  ferment  appears  in  seeds  from  the  commence- 
ment of  germination  ;  this  takes  place  in  the  potato  in 


SOLUBLE  FERMENTS. 


287 


spring  ;  then  the  fermentiferous  agent  shows  itself  in  the 
tubercle,  as  it  did  in  the  germinated  barley  ;  it  liquefies 
the  starch,  and  puts  it  into  a  condition  to  be  distributed 
over  the  parts  where  it  may  minister  to  the  nutrition, 
that  is  to  say,  to  the  development  of  the  life  of  the 
plant. 

"  In  most  animals,  the  phase  of  the  production  of 
glycogen,  and  that  of  its  fermentation,  are  not  so  distinc 
as  in  plants.  The  two  phenomena  are  often  continuous 
and  simultaneous.  An  exception  must,  however,  be 
made  with  respect  to  the  earliest  periods  of  life, 
especially  in  those  animals  which  undergo  metamor- 
phoses. For  example,  if  we  consider  the  larva  of  the 
common  fly,  miisca  hccelia,  the  gentle — to  call  it  by  its 
common  name — we  find  that  it  contains  an  enormous 
quantity  of  starch.  It  is  truly  a  bag  full  of  glycogen. 
At  this  time  we  find  nothing  else  but  glycogen,  and  not 
even  a  trace  of  sugar.  The  reason  of  this  is  that  the 
glycose  ferment  does  not  yet  exist.  But  soon  the 
chrysalis  will  succeed  to  the  larva,  and  then  in  the  new 
phase  of  existence,  during  which  the  perfect  animal  is 
in  formation,  the  reserve  of  glycogen  must  be  utilized. 
The  ferment  appears,  and  the  starch  is  liquefied. 

Something  analogous  to  this  is  seen  in  beings  of  a 
higher  order,  for  example,  among  the  mammalia,  at 
that  period  of  the  embryonic  life  when  nutrition  is 
hastened,  and  the  plastic  and  formative  activity  attains 
its  highest  degree.  The  glycogen  matter  deposited  at 
various  points  of  the  foetus,  and  of  its  envelopes,  is  set 
in  motion,  dissolved,  and  transformed  into  sugar.  * 

*  The  experiments  of  CI.  Bernard,  Hensen,  Magendie,  and  Schiff  prove  that 
in  living  blood  there  exists  a  soluble  ferment,  capable  of  transforming  starch 


288 


ON  FERMENTATION. 


The  digestion  of  starches  consists  in  their  trans- 
formation into  soluble  and  assimilable  substances,  solu- 
ble in  order  to  be  able  to  circulate  from  one  part  of 
the  organism  to  another.  Digestion  is,  therefore,  the 
prologue  of  the  act  of  nutrition.  Whenever  these 
starchy  matters  are  to  nourish  an  organism,  we  shall 
find  this  previous  preparation.  But  all  organisms,  in 
the  vegetable  as  well  as  in  the  animal  kingdom,  employ 
starchy  substances  as  food,  all,  therefore,  digest  these 
substances,  in  the  strict  sense  of  the  word." 

The  agents  in  these  acts  of  digestion  are  the  diastase 
of  germinated  barley,  saliva,  pancreatic  juice,  &c. 

The  action  of  diastase  on  starch  is  exerted  at  the 
ordinary  temperature,  and  attains  its  maximum  effect 
at  75°  C.  (167°  R).  Boiling  destroys  the  specific  power 
of  this  substance. 

It  is  almost  useless  to  mention,  since  the  fact  is  so 
well  known,  that  the  eff*ects  of  diastase  on  starch  are 
imitated  by  the  intervention  of  boiling  dilute  sulphuric 
acid. 

Alterative  Ferment. — Cane,  sugar,  or  saccharose,  C^'"* 
j^22  Qii^  transformed,  as  is  proved  by  the  researches 
of  M.  Dubrunfant  (Ann.  de  Chim.  et  de  Phys.  (3), 
vol.  21,  169,  1847  j  Comp.  Rend,  de  TAcad.  vol.  29,  p.  51, 
1849,  and  vol.  42,  p.  901,  1856)  by  becoming  hydrated, 
under  the  influence  of  dilute  mineral  and  even  organic 

and  glycogen  into  sugar.  This  ferment  is  also  found  in  very  fresh  liver.  It 
may  be  eliminated  from  it  by  making  a  cold  infusion  of  a  liver,  which,  on 
account  of  general  disease,  produces  no  more  sugar,  and  contains  no  glycogen, 
and  by  precipitating  it  subsequently  by  alcohol.  The  deposit,  redissolvcd  in 
water,  acts  on  starch.  This  special  ferment  exists  in  the  blood  of  frogs  in  spring 
and  in  summer.  In  fact,  dextrin  injected  into  their  veins  passes  with  the  urine 
in  the  form  of  sugar.  In  winter  it  is  wanting,  for  dextrin  appears  in  the  urine 
without  alteration. 


SOLUBLE  FERMENTS. 


289 


acids,  into  sugar,  which  causes  the  plane  of  polarization 
to  turn  to  the  left,  and  energetically  reduces  Fehling's 
cupro-potassic  reagent.  Saccharose,  on  the  contrary, 
turns  the  plane  of  polarization  to  the  right,  and  has 
no  action  on  Fehling's  liquid.  The  alteration  is  pro- 
duced in  the  cold,  but  more  rapidly  in  the  warm  pro- 
cess ;  it  has  been  effected  by  simply  boiling  solutions 
of  cane-sugar  in  water,  by  their  exposure  to  light,  or 
by  the  mechanical  means.  Thus  the  mere  pulverizing 
the  sugar  causes  a  small  portion  of  altered  sugar  to  be 
formed.  The  product  of  the  alteration  is  not  a  unique 
and  simple  principle. 

The  same  chemist  proved  that  saccharose,  after 
alteration,  is  found  to  be  changed  into  two  sugars  of 
the  formula  H^^  one  of  which  turns  the  plane  of 
polarized  light  to  the  right,  and  is  identical  with  grape- 
sugar,  or  ordinary  glucose ;  the  other,  on  the  contrary, 
causes  the  plane  of  polarization  to  deviate  to  the  left ; 
this  is  the  uncrystallizable  sugar  of  acid  fruits,  or 
Icevtdose, 

As  at  15°  C.  (59°  F.),  the  specific  rotatory  power  of 
glucose  is  equal  to  -f-  57*8°,  whilst  that  of  laevulose 
is  —  106°,  and  as  the  two  sugars  are  found  in  the  mix- 
ture in  equal  proportions  by  weight,  it  can  be  under- 
stood that  this  mixture,  called  altered  sugar,  has  a 
laevo-rotatory  action  due  to  the  excess  of  one  of  these 
powers  over  the  other. 

Altered  sugar  has  a  laevo-rotatory  power  of  25*"  at 
the  temperature  of  15°  C. 

The  formation  of  these  two  sugars  (glucose  and 
laivulose)  is  easily  formulated  by  the  following  equa- 
tion : — 


290 


ON  FERMENTATION. 


QI2  ll^i  QM   ^  O   =  H^2  Q6  ^  c« 

Saccharose.        Water.         Glucose.  Loevulose. 

This  is  confirmed  by  the  fact,  that  under  the  influence 
of  heat,  saccharose  is  changed  into  glucose  and  laevu- 
losan  (laevulose  less  water),  which  is  itself  changed  into 
tevulose  under  the  influence  of  acids. 

The  following  is  the  manner  in  which  M.  Dubrunfant 
brings  about  the  separation  of  these  two  bodies. 

Ten  grammes  of  altered  sugar  are  dissolved  in  100 
grammes  of  water,*  and  the  whole  heated  with  six 
grammes  of  calcium  hydrate.  The  mixture,  which  at 
first  is  fluid,  forms  into  a  mass  at  the  end  of  a  short 
time ;  this  is  the  calcium  laevulosate,  which  separates  as 
it  crystallizes,  while  the  glucosate  remains  in  the  mother- 
liquor.  The  mass  is  pressed  at  once,  and  the  solid 
and  liquid  parts,  after  having  been  dissolved  or  diluted 
with  a  suitable  quantity  of  water,  are  decomposed 
separately  by  carbon  dioxide ;  the  former  furnishes  a 
solution  of  tevulose,  the  latter  of  glucose.  We  can  also 
efi*ect  a  separation,  or  at  least  isolate  the  laevulose 
free  from  glucose,  by  stopping  a  fermentation  of  cane- 
sugar  almost  in  the  middle  of  its  course.  The  glucose 
fermenting  more  easily  than  the  laevulose,  the  latter  is 
found  alone. 

Without  dwelling  any  further  on  all  these  proofs  of  a 
true  splitting  up,  which  leave  no  doubt  as  to  the  nature 
of  the  reaction,  let  us  proceed  to  the  ferment. 

This  alteration,  this  splitting  up  of  the  saccharose  by 

*  According  to  the  experiments  made  at  the  laboratory  of  the  Sorbonne,  the 
proportion  of  water  indicated  by  Dubrunfant  is  too  great  to  allow  of  the  crys- 
tallization of  the  calcium  laevulosate.  Better  results  were  obtained  by  em- 
ploying four  parts  of  water  to  one  part  of  altered  sugar. 


SOLUBLE  FERMENTS. 


hydratation  is  produced  before  any  alcoholic  fermenta- 
tion takes  place.  Cane-sugar  can  not  be  resolved  into 
alcohol  and  carbon  dioxide  till  after  it  has  been  altered, 
but  beer-yeast  itself  performs  this  operation.  The 
action  was,  at  first,  attributed  to  the  acidity  of  the 
yeast,  and  M.  Pasteur  thought  that  it  was  the  result  of 
the  presence  of  succinic  acid. 

M.  Berthelot  was  the  first  to  show  that  the  alteration 
of  cane-sugar  by  yeast  is  independent  of  the  condi- 
tions of  acidity  and  neutrality ;  that  it  is  due  to  the 
intervention  of  a  special  soluble  ferment  contained  in 
the  cells  of  yeast.  This  principle  is  found  in  the  water 
in  which  yeast  has  been  washed,  and  acts  energetically 
upon  sugar,  even  in  a  neutral  liquid. 

The  properties  of  this  soluble  ferment,  besides  its 
peculiar  activity,  cause  it  to  resemble  all  the  bodies  of 
this  class.  I  have  ascertained  that  it  is  easy  to  sepa- 
rate it  entirely  from  the  albuminoid  substances  which 
accompany  it,  by  following  the  method  of  precipitation 
by  phosphoric  acid  and  lime-water  mentioned  above.* 

Saccharose  is  not  susceptible  of  being  absorbed  and 
assimilated  under  its  original  form,  any  more  than  amy- 
laceous matter,  although  it  has  the  advantage  over  the 
latter  in  its  solubility.  Like  starch,  it  has  to  be  digested, 
and  the  products  of  this  digestion  are  glucose  and  laevu- 
lose.-f-     M.  Bernard  has  proved  the  existence  in  the 

*  M.  Bechamp  found,  after  M.  Berthelot,  the  alterative  ferment  (zymase)  in 
the  water  of  yeast,  spontaneously  weakened  by  want  of  nourishment.  Under 
these  conditions,  I  have  myself  observed  that  a  liquid  is  obtained,  excessively 
active,  and  which  produces  alteration  almost  instantaneously. 

f  Cane-sugar  is,  according  to  the  expression  of  M.  Bernard,  like  an  inert  or 
indifferent  matter  which  can  circulate  with  impunity  in  the  blood  or  the  sap, 
without  the  anatomical  elements  being  able  to  turn  it  aside  or  to  appropriate  it. 
He  gives  as  a  proof  of  this,  that  when  cane-sugar  is  injected  into  the  veins  or 


J92 


ON  FERMENTATION. 


intestinal  juice  of  a  soluble  alterative  ferment,  similar 
to  that  of  yeast ;  it  forms  one  of  the  most  important 
and  useful  elements  of  this  secretion.  In  order  to  prove 
this,  a  solution  of  pure  saccharose  (not  reducing  Fehling's 
liquid)  must  be  injected  into  a  portion  of  the  intestine 
included  between  two  ligatures,  or  placed  in  contact 
with  an  infusion  of  intestinal  mucous  membrane  ;  we 
then  see,  in  a  very  short  time,  that  the  sugar  reduces  the 
copper  oxide.  It  may  be  ascertained  by  the  saccha- 
rometer  that  the  deviation  of  this  physiologically  altered 
sugar  is  the  same  as  that  of  chemically  altered  sugar. 
M.  CI.  Bernard  has  ascertained  the  existence  of  altera- 
tive ferment  in  dogs,  rabbits,  birds,  and  frogs.  M.  Bal- 
biani  has  proved  that  it  is  to  be  found  in  the  digestive 
tube  of  silkworms.  M.  Bernard,  by  comparing  these  facts 
relating  to  the  digestion  of  saccharose  in  animals,  with 
the  analogous  transformations  observed  in  the  vegetable 
economy,  as  he  had  before  done  for  the  saccharification 
of  starch,  shows  that  the  alterative  ferment  is  generally 
found  in  all  parts  of  the  living  economy,  and  under  all 
the  circumstances  in  which  saccharose  is  necessary  for 
nutrition.  Sugar-cane  which  is  flowering,  and  beet  which 
runs  to  seed,  transform  by  alteration  the  sugar  laid  up  in 
their  tissues.  The  agent  is  always  exactly  the  same,  an 
alterative  ferment.  M.  Bernard  obtained  it  from  beet- 
root in  course  of  evolution,  as  M.  Berthelot  had  extracted 
it  from  yeast  Alcoholic  fermentation,"  says  our  illus- 
trious physiologist,  is  a  phenomenon  correlative  to  the 
nutrition  of  an  organism,  the  yeast  of  beer,  torula 

cellular  tissue  of  an  animal,  it  is  returned,  weight  for  weight,  in  the  urinary 
excretion,  and  consequently  passes  through  the  economy,  without  being  modi- 
fied or  assimilated. 


1 


SOLUBLE  FERMENTS. 


cerevisicB.  But  saccharose  is  unfit  for  the  nutrition  of  this 
microscopic  form  of  hfe,  as  it  is  unsuited  to  that  of 
higher  forms.  It  is  therefore  necessary  that  saccharose 
should  be  modified,  and  transformed  into  glucose,  before 
it  can  conduce  to  the  vital  changes  of  the  organic 
ferment.  The  cell  of  yeast,  by  producing  this  trans- 
formation, tends  to  its  own  development.  It  digests  the 
saccharose  for  itself ;  alteration  is  here  once  more  a 
digestive  action  of  the  same  nature  as  those  which  we 
have  already  examined. 

"  In  the  same  manner  as  starch,  the  saccharose,  which 
exists  in  a  state  of  reserve  in  the  tissues  of  a  great 
number  of  plants,  is  unfit  to  participate  in  the  nutritive 
circulation  of  the  plant.  It  is  for  this  reason  tliat  sugar 
can  accumulate  and  be  laid  up  in  store,  as  occurs  in  the 
root  of  the  beet,  and  in  the  stem  of  the  sugar-cane. 
The  sugar  forms  a  reserve  there,  while  it  waits  for  the 
moment  to  begin  to  act.  This  period  arrives  for  beet- 
root, when  it  is  about  to  put  forth  buds,  flowers,  and 
fruit ;  then  the  sugar  diminishes  by  degrees,  and  disap- 
pears soon  after  from  the  tissue  and  the  stalk  of  the 
beet,  changing  into  glucose.  The  leaves  at  this  time 
contain  glucose  only  ;  the  root  yields  up  its  store,  and 
the  reserves  of  sugar  which  it  contained  are  distributed 
through  the  stalk  to  serve  for  the  purposes  of  the  flower 
and  seed  ;  but  this  is  not  possible,  except  by  means  of  a 
previous  transformation,  which  changes  the  chemical 
nature  and  the  composition  of  the  saccharose  and  causes 
it  to  pass  into  the  state  of  glucose.  This  is  again  a  true 
digestion.  Beet,  therefore,  like  yeast,  and  like  animals, 
must  digest  its  sugar." 

I  could  not  resist  the  pleasure  of  giving  the  very  words 


294 


ON  FERMENTATION. 


of  our  great  observer.  Nothing  can,  in  fact,  give  a 
clearer  idea  of  the  importance  of  the  subject  of  which 
we  are  now  treating,  than  the  wide  scope  of  the  con- 
siderations by  which  these  phenomena  are  connected 
with  the  entirety  of  the  great  act  of  nutrition  in  Hving 
organisms.  We  have  not  here  to  deal  with  isolated 
chemical  reactions,  interesting  on  account  of  the  obscu- 
rity which  still  reigns  over  the  cause  which  produces 
them,  but  with  transformations  which  play  an  important 
part  in  the  acts  of  life  and  nutrition,  the  remarkable 
generality  of  which  is  placed  in  the  most  striking  point 
of  view. 

Eimdsive  and  Saponifying  Ferment. — M.  CI.  Bernard 
has  shown  that  the  pancreatic  juice  is  the  only  one 
among  all  the  liquids  secreted  in  the  digestive  tube, 
which  possesses  the  remarkable  property  of  forming  an 
emulsion,  and  afterwards  saponifying  neutral  fatty  sub- 
stances. It  constitutes,  therefore,  the  true  agent  of 
digestion  for  fatty  substances  or  natural  glycerids. 

The  emulsion  which  precedes  saponification  is  rather 
a  physical  than  a  chemical  transformation.  It  is  the 
mechanical  division  of  the  fatty  liquid  which  is  thus 
separated  into  an  infinite  number  of  little  globules, 
maintaining  themselves  by  means  of  a  characteristic 
molecular  constitution.  Under  the  microscope,  we  see  a 
great  number  of  granular  bodies  swimming  in  the  liquid, 
and  animated  by  the  Brownian  movement.  Emulsion 
is  the  necessary  condition  for  the  absorption  of  fatty 
bodies. 

Among  the  organic  secretions  of  the  digestive  canal, 
the  pancreatic  juice  is  the  only  one  which  can  furnish 
with  oils  a   complete   and   persistent  emulsion.  M. 


SOLUBLE  FERMENTS. 


29s 


Bernard  attributes  this  emulsive  power  to  a  peculiar 
soluble  ferment.  He  considers  that  the  persistent  emul- 
sion of  the  fat  in  milk  is  due  to  the  presence  of  an 
emulsive  ferment,  and  that  this  nutritive  liquid  contains 
the  fatty  matter  ready  prepared  for  absorption. 

Emulsion  by  the  pancreatic  juice  or  the  extract  of  the 
gland  is  instantaneous.  A  slower  action  ensues  ;  this  is 
saponification,  that  is  to  say,  resolution  of  trimargarin, 
triolein,  tristearin,  &c.,  by  hydratation,  into  glycerin  and 
fatty  acids.  This  resolution  is  not  always  effected  in  the 
intestine.  The  emulsion  penetrates  into  the  chyliferous 
vessels  still  retaining  its  lactescent  character,  and  it  is 
not  till  afterwards,  in  the  different  tissues,  that  saponifi- 
cation takes  place.  It  has  been  thought  that  the  saponi- 
fication excited  by  the  pancreatic  juice  was  due  to  the 
alkalinity  of  this  liquid.  M.  Bernard  replies  to  this 
objection  by  showing:  (i)  That  other  secretions  quite 
as  alkaline  are  inactive  ;  (2)  That  the  tissue  of  the 
pancreas,  which  has  no  alkaline  reaction,  rapidly  pro- 
duces phenomena  of  the  same  kind  ;  (3)  We  can 
destroy  by  boiling,  the  specific  power-of  the  juice,  without 
weakening  its  alkalinity.  The  emulsive  ferment  may  be 
eliminated,  mixed  with  albuminoid  matter,  and  two  other 
indirect  ferments,  of  which  we  have  already  spoken  (pan- 
creatic diastase,  and  the  digestive  ferment  of  the  albu- 
minoids). The  emulsive  ferment  possesses  the  property 
of  casein,  of  being  precipitated,  in  the  cold,  by  magne- 
sium sulphate.    It  is  coagulated  by  heat. 

We  find  bodies  of  the  same  order  in  plants.  If  we 
steep  oleaginous  seeds  in  water,  we  obtain  an  emulsion 
in  which  we  soon  find  glycerin  and  the  free  fatty  acids. 
During  the  germination,  the  emulsive  and  saponifying 


296 


ON  FERMENTATION. 


ferment,  placed  with  fat  in  the  presence  of  water,  causes 
it  to  undergo  true  digestion,  and  renders  it  assimilable. 

The  emulsine  of  almonds  has  an  analogous  property ; 
we  shall  presently  see  that  it  acts  on  certain  glucosides. 
We  cannot  determine  whether  this  complicated  action 
depends  on  a  mixture  of  several  ferments,  or  on  a  vari- 
able activity  of  the  same  body,  according  to  the  elements 
subjected  to  it.  Kolliker  and  Miiller  have  shown  that 
the  pancreatic  juice  can  effect  the  decomposition  of 
amygdalin.    This  is  a  very  important  indication. 

Perhaps  we  shall  find,  when  we  have  succeeded  in 
isolating  the  emulsive  ferment  of  the  pancreatic  juice, 
that  it  has  the  same  properties  as  emulsin*  or  synastase. 

As  a  conclusion  of  that  which  relates  to  the  fermenta- 
tion of  glycerides,  we  have  only  now  to  obtain  definite 
ideas  as  to  the  kind  of  reactions  which  take  place,  by 
giving  the  equations  of  the  decompositions,  as  the  result 
of  the  labours  of  M.  Berthelot  has  shown  them. 

1.  Emulsion  ;  a  physical  change. 

2.  Splitting  up. 

H«  (C®  H^^  O  .  O)'  +  3       O  =  O*^  K8  Qs  +  3  (C's  H-^=^  O .  HO) 
Triolein.  Water.  Glycerin.  Oleic  acid. 

C3       (C^8        O  .  0;3  +  3  H2  O  r=  C3  H8  O^  +  3  {C^  H35  O  .  HO) 
Tristearine.  Glycerin.  Stearic  acids. 

C3  H^       H7  O  .  0)«  +  3  H'^  O  =      H8  O'''  +  3  (C^  H^  O  .  HO) 
Tributyrin.  Glycerin.  Butyric  acid. 

Albinniuosic  Fervicnt. — Two  digestive  secretions,  the 
gastric  and  the  pancreatic  juices,  and  probably  the 
intestinal  juice  also,  possess  the  power  of  transforming 
the  soluble  or  insoluble  but  indiffusible  albuminoid 
substances  into  soluble  and  diffusible  principles.  It 


SOLUBLE  FERMENTS. 


297 


has  been  found  that  this  property  is  due,  as  is  the  case 
with  the  digestion  of  starchy  bodies  and  saccharose,  to 
the  intervention  of  special  nitrogenous  principles,  to 
which  the  name  of  pepsin  has  been  given  (the  digestive 
principle  of  the  gastric  juice),  and  to  that  of  the  albu- 
minosic  ferment  of  the  pancreatic  juice. 

Pepsin,  the  method  of  extracting  which  we  know 
(processes  of  Schwann,  Wasmann  and  Vogel,  and  de 
Briicke),  only  acts  on  albuminoids  in  an  acid  medium, 
and  within  very  restricted  limits  of  temperature.  Its 
action  is  principally  exerted  on  fibrin.  The  short  time 
that  food  continues  in  the  stomach  only  allows,  in  any 
case,  very  slight  transformations  ;  and  we  must,  indeed, 
admit,  that  the  stomachic,  acid,  pepsinic  digestion  of 
albuminoid  substances  is,  in  every  respect,  a  very  incom- 
plete operation,  and  rather  preparatory  than  final. 

Notwithstanding  all  that  has  been  written  and  said 
upon  the  chemical  question,  but  very  little  is  yet  known 
about  the  real  transformations  of  the  albuminoids  in  the 
stomach,  or  under  the  influence  of  artificial  gastric  juice  ; 
we  will  not,  therefore,  enter  into  long^details  on  this  sub- 
ject ;  they  would  only  give  us  superabundant  proofs  that 
everything  remains  to  be  done.  In  fact,  until  the  con- 
stitution of  albuminoids  is  definitely  ascertained,  and  we 
know  all  the  intermediate  terms  of  their  transformations 
and  progressive  splittings  up,  we  shall  not  be  able  to  enter 
on  this  question  with  advantage  and  success.  I  shall 
therefore  content  myself  with  a  few  observations. 

Action  of  the  Gastric  J  idee  on  the  Fibrin  of  Blood, — 
According  to  Briicke,  uncooked  fibrin,  placed  in  contact 
at  40""  C.  (104°  F.)  with  a  sufficient  quantity  of  gastric 
juice  of  good  quality,  swells  and  dissolves  rapidly, 


298 


ON  FERMENTATION. 


changing  at  first  into  a  principle  soluble  in  dilute  acids, 
but  capable  of  being  precipitated  by  neutralization  ;  this 
principle  seems  to  have  all  the  characters  of  syntonin, 
the  product  of  the  transformation  of  muscular  tissue 
under  the  influence  of  acids.  By  prolonged  digestion, 
the  syntonin  disappears  in  its  turn,  and  gives  place  to  a 
unique  principle,  peptone.  It  is  doubtful  whether  peptone 
has  time  to  form  in  the  stomach. 

Meissner,  on  the  contrary,  asserts  that  the  syntonin, 
which  originates  at  first,  splits  up  by  an  ulterior  action 
into  an  insoluble  indigestible  substance,  parapeptone,  not 
capable  of  being  subsequently  transformed  by  the  gastric 
juice,  and  into  soluble  principles  (peptones  a  b  c).  The 
acid  stomachic  digestion  seems,  according  to  these  views, 
to  be  a  decomposition  of  proteinic  substances.  This 
manner  of  considering  the  question  appears  to  me  the 
most  probable,  and  the  most  conformable  to  the  analo- 
gies that  we  can  find,  according  to  what  is  known  of  the 
reactions  of  the  albuminoids. 

Other  albuminoid  substances  behave,  according  to 
Meissner,  nearly  in  the  same  manner  as  fibrin,  by  be- 
coming previously  changed  into  syntonin. 

The  digestive  action  of  the  pancreatic  juice,  with 
reference  to  albuminoids,  is  much  more  energetic  and 
efficacious  than  that  of  pepsin.  It  does  not  require 
such  limited  conditions.  Thus  the  albuminosic  ferment 
of  the  pancreas  develops  its  power  as  well  in  an  acid  as 
in  an  alkaline  liquid.  The  products  formed  are  distin- 
guished from  the  initial  compounds  by  the  property  of 
being  no  longer  precipitated  by  the  neutralization  of  the 
liquid  ;  they  are  incoagulable  and  easily  diffusible,  thus 
resembling  crystalloids.    Without  wishing  to  speak  very 


SOLUBLE  FERMENTS.  299 

decidedly  on  this  question,  I  am  inclined  to  consider 
peptones  as  bodies  closely  allied  to  the  diffusable,  soluble, 
uncrystallizable,and  incoagulable  substances  which  I  have 
obtained  by  the  incomplete  action  of  hydrated  baryta  on 
albumin.  As  to  the  characters  of  the  third  order,  by 
which  an  attempt  has  been  made  to  distinguish  the  dif- 
ferent peptones,  we  pass  them  over  in  silence ;  they 
present  no  definite  characters  to  the  mind.  We  can 
form  a  clear  idea  of  the  nature  of  these  bodies  only  by 
bringing  about  by  hydratation  a  complete  resolution  of 
these  bodies  into  definite  chemical  principles,  such  as 
the  elements  of  urea,  tyrosine,  leucine,  and  its  homolo- 
gues,  and  thus  determining  their  constitution.  An  in- 
vestigation of  this  kind,  connected  with  that  which  I  have 
undertaken  with  respect  to  albuminoid  substances,  could 
not  fail  to  produce  important  results  in  a  physiological 
point  of  view. 

The  analogy  of  functions  led  M.  Bernard  to  seek  for 
albuminosic  digestion  in  plants.  If,  with  him,  we  call  by 
this  name  every  transformation  of  albuminoid  matter 
into  soluble  diffusible  principles,  it  is  certain  that  it  must 
exist  there. 

Thus  the  phenomena  presented  by  yeast  preserved  in 
a  damp  state  without  nourishment,  and  to  which  we  have 
referred  when  treating  of  alcoholic  fermentation,  ought 
and  must  be  considered  as  a  true  digestion  of  proteids. 
It  is  the  same  with  the  continuous  chemical  transforma- 
tions which  take  place  in  the  protoplasm  of  vegetable 
and  animal  cells.  Nothing  proves  that  the  first  products 
of  the  change  of  albuminoids  in  the  organism  are  entirely 
excrementitious  bodies,  and  that  these  products,  as  long 
as  they  remain  above  a  certain  limit  of  splitting  up, 


300 


ON  FERMENTATION. 


which  they  do  not  attain,  as,  for  example,  the  crystal- 
loids, such  as  leucine,  tyrosine,  &c.,  can  no  longer  con- 
tribute to  organic  synthesis,  and  to  the  formation  of  new 
cells  or  tissues.  It  is  the  water  with  which  digested 
yeast  has  been  washed  that  contains  the  most  suitable 
nourishment  for  the  development  of  this  organic  ferment ; 
but,  excepting  the  albumin,  which  is  inactive  as  nourish- 
ment for  yeast,  the  nitrogenous  principles  of  the  water 
of  yeast  are  products  of  an  order  inferior  to  that  of 
proteinic  substances ;  they  are  the  results  of  their 
splitting  up.  We  do  not  here  speak  of  leucine  and 
tyrosine,  the  presence  of  which  has  been  recognized  in 
the  water  with  which  yeast  has  been  washed,  and  which 
have  no  nutritive  power,  but  only  of  the  nitrogenous 
bodies  contained  in  the  uncrystallizable  sirup. 

Briicke  found  pepsin  in  the  blood,  the  muscles,  and 
the  urine.  This  agent,  therefore,  is  not  confined  to  the 
stomach  only  ;  it  is  diffused  over  the  various  parts  of 
the  organism,  wherever  its  presence  may  be  necessary, 
especially  where  there  are  albuminoids  to  be  liquefied 
and  digested  in  order  to  render  them  fit  for  nutrition. 
Bretonneau  had  already  announced  that  meat  intro- 
duced into  a  sub-cutaneous  wound  could  be  digested 
there  as  in  the  stomach. 

We  have  given  these  details  of  the  indirect  fermen- 
tations which  are  connected  with  the  great  biological 
phenomena  of  digestion,  the  first  act  of  nutrition  in 
living  organisms,  because  of  the  great  importance  that 
attaches  to  this  subject. 

As  to  the  other  phenomena  of  the  same  order,  wc 
shall  treat  them  more  slightly,  since  all  that  is  essential 
may  be  described  in  general  terms. 


SOLUBLE  FERMENTS. 


301 


Fermentation  of  Gliccosides, — The  ferment  which  acts 
most  generally  and  energetically  on  these  bodies  is  found 
in  sweet  and  bitter  almonds ;  this  is  synastase  or 
emulsin.  It  acts  best  between  30""  and  40°  C.  (86° 
and  104°  F.)  ;  but  still  it  can  bear  an  elevation  of 
temperature  to  80°  C.  (176°  F.),  without  losing  its 
specific  power. 

Acids  and  alkalies,  in  small  quantities,  do  not  inter- 
fere with  the  action  of  emulsin.  It  is  only  after  having 
undergone  a  somewhat  advanced  state  of  putrefaction 
that  emulsin  loses  its  qualities.  Its  intervention  may 
be  supplied,  up  to  a  certain  point,  by  other  principles 
of  animal  origin.  Substances  which  are  poisonous  to 
plants  and  to  yeast  have  no  influence  on  the  reactions 
produced  by  synastase. 

All  these  facts,  in  addition  to  its  mode  of  preparation 
{see  the  process  of  Wittich,  before  described),  leave  no 
doubt  as  to  the  nature  of  emulsin  ;  it  is  a  soluble  fer- 
ment of  the  diastase  family. 

It  will  be  sufiicjent  to  give  a  short  summary  of  the 
reactions  over  which  it  presides  ;  for  further  details  w^e 
must  refer  to  works  on  chemistry. 

1st.  Splitting  up  of  amygdalin  into  glucose,  hydro- 
cianic  acid,  and  benzoyl  hydride.  This  reaction  ex- 
plains that  which  takes  place  when  bitter  almonds  are 
pounded  with  water.  The  bitter  almond  contains  at 
the  same  time  amygdalin  and  emulsin.  These  two 
bodies  are  in  distinct  cells,  and  can  only  react  on  each 
other  when  a  mechanical  action  and  solution  place 
them  in  close  contact. 

This  action  may  take  place  within  the  organism  ;  as, 
for  instance,  when  we  inject  dissolved  amygdalin  and 


302 


ON  FERMENTATION. 


emulsin  into  the  veins  of  an  animal,  the  subject 
dies  with  the  symptoms  of  poisoning  by  prussic 
acid. 

2nd.  Resolution  of  salicin  into  glucose  and  saligenin. 
3rd.        „         of  chlorosalicin    into    glucose  and 

chlorosaligenin. 
4th.         „         of  helicin  into  glucose  and  salicylic 

hydride. 

5th.        „        of  arbutin  into  glucose  and  hydro- 
quinone. 

6th.        „         of  phlorizin  into  glucose  and  phlo- 
retin. 

7th.         „        of  esculin  into  glucose  and  esculetin. 
8th.        „         of  daphnin  into  glucose  .and  daph- 
netin.* 

Vanillin,  the  odorous  principle  of  vanilla,  has  quite 
recently  been  artificially  produced,  by  decomposing  by 

*  These  decompositions  take  place  according  to  the  following  equations: — 

C^o       N  Oi»  +  2      O  =  2  06  4.  a  H«  O  +  C  N  H 

Amygdalin.  Water.  Glucose.  Benzoyl  Prussic 

hydride.  acid. 

Ql^  H'«  07   +    H2  O    =    C6  Q6    ^    Q7  Q2 

Salicin.  Glucose.  Saligenin. 

C'^       ci.  07  +  H2  O  =      H^^^  0«  +  a      CI.  O^ 

Chlorosalicin.  Glucose.  Chlorosaligenin. 

Cisf^ieo^   J.  H'^o  =  C^Hi^QG    ^  C7H«0- 

Helicin.  Glucose.  Salicylic  hydride. 

Qu       07  +  H2  O  =      H^^  0«  4-  O^ 

Arbutin.  Glucose.  Hydroquinone. 

C2«        O'^  +  H2  O  =  a  H'*-*  O^  4-  O^ 

Phlorizin.  Glucose.  Phloretin. 
Q2i  ii'24  o^'  -h  3  H2  O  =  2  C«  H'*^  G«  +  C»  H«  O* 

Esculin.  Glucose.  Esculetin. 

QV         o«9  +  2  H2  O  =  2  C«  H^'^  O'''  +  C»  09 

Dai^hnin.  Glucose.  Daphnetin. 


SOLUBLE  FERMENTS. 


303 


means  of  synoptase  a  glucoside  contained  in  coniferous 
plants  (coniferin),  and  by  oxidizing  the  principle  formed 
by  this  decomposition. 

Emulsin,  like  most  of  the  soluble  ferments,  may  be 
replaced,  with  respect  to  glucosides,  by  agents  purely 
chemical.  Thus  we  arrive  at  the  same  results,  in  most 
cases,  by  boiling  with  dilute  acids. 

A  very  interesting  chemical  reaction,  of  the  same 
order  as  that  which  we  have  observed  in  bitter  almonds, 
is  found  in  the  flour  of  mustard  seed,  mixed  with  water. 
This  mixture,  the  common  mustard  plaster,  has,  as  we 
know,  a  strong  odour  and  a  burning  taste.  The  proper- 
ties of  this  sinapism  are  due  especially  to  the  presence 
of  essence  of  mustard,  or  allyl  sulphocyanide.  Eut  this 
essence  does  not  exist  fully  formed  in  the  seed  any  more 
than  the  essence  of  bitter  almonds  in  the  kernel  ;  it 
originates  at  the  expense  of  a  special  compound 
contained  in  black  mustard,  under  the  influence  of 
a  soluble  ferment,  myrosin,  contained  in  white  or 
black  mustard,  and  generally  in  the  seeds  of  the 
cruciferae. 

The  initial  product  has  been  isolated  in  a  crystalline 
form  ;  this  is  the  potassic  salt  of  a  complex  acid,  myronic 
acid.  As  to  the  ferment,  we  have  nothing  of  importance 
to  say  of  it ;  it  may  be  eliminated  from  the  seeds  of  the 
cruciferae,  or  of  white  mustard,  which  do  not  contain 
potassium  myronate,  by  the  general  methods  already 
described  ;  its  physical  characters,  as  well  as  its  com- 
position, are  those  of  all  the  soluble  ferments. 

Potassium  myronate  H*'  N  .  K.  S'  O'*'),  whose 
presence  characterizes  black  mustard,  is  resolved,  under 
the  influence  of  myrosin,  into  glucose,  allyl  sulphocyanide, 


304 


ON  FERMENTATION. 


and  potassium,  bisulphate,  as  shown  in  the  following 
equation  : — 

CO  Ills  N  K  S2  O^o  =  C«  H^-  0<5  +  C  N  C3      I  S  +  S  O*  K  H 
Potassium  myronate.        Glucose.  AUyl  sul-  Potassium 

phocyanide.  bisulphate. 

The  potassium  myronate  is  extracted  in  the  following 
manner  by  the  process  of  M.  Bussy,  modified  by 
Messrs.  Witt  and  Korner.  A  kilogramme  of  black 
mustard  seed  pulverized  is  boiled  with  litres  of 

alcohol  at  82  per  cent,  until  a  quarter  of  a  litre  of 
alcohol  is  distilled  over.  It  is  pressed  while  hot,  and 
the  same  operation  is  repeated  with  the  remainder. 
This  is  again  pressed,  dried  at  100°  C.  (212°  F.),  pul- 
verized, and  digested  for  twelve  hours  with  three  parts 
of  cold  water ;  it  is  pressed  again,  and  the  remainder 
diffused  in  two  parts  of  cold  water.  The  watery  solu- 
tions are  evaporated,  after  the  addition  of  a  little  barium 
carbonate,  till  they  are  of  a  sirupy  consistence.  The 
remainder  is  treated  with  boiling  alcohol  at  85  per  cent, 
(i  or  litres)  ;  it  is  then  filtered,  distilled,  and  left  in 
flat  open  plates  to  crystallize. 

We  will  now  mention  the  other  analogous  fermenta- 
tions, without  dwelling  on  them. 

I.  The  fresh  root  of  the  madder  contains  alizarin  and 
other  colouring  matters,  insoluble  of  themselves,  under 
the  form  of  soluble  glucosides  (Rubi.an,  ruberythric  acid). 
Together  with  these  principles,  the  same  root  contains 
a  soluble  ferment,  erythrozyme,  which  is  not  slow  in 
splitting  up  these  glucosides,  when  the  powdered  mad- 
der-root is  mixed  with  water.  Under  its  influence, 
the  colouring  matters,  at  first  dissolved,  separate  whilst 


SOLUBLE  FERMENTS. 


305 


sugar  is  formed.  M.  E.  Kopp  has  succeeded  in  stopping 
the  action  of  the  ferment,  by  the  addition  of  a  certain 
quantity  of  sulphurous  acid  ;  he  founded  on  this  fact  a 
very  interesting  process  for  the  extraction  of  the  colour- 
ing matters  of  madder. 

2.  The  infusions  of  the  seeds  of  buckthorn,  when  they 
are  kept  for  some  time,  also  undergo  a  fermentation 
which  decomposes  the  soluble  colouring  glucosides  con- 
tained in  this  infusion  or  decoction  ;  glucose  and  an 
insoluble  pigment  are  formed.  Boiling  with  acids  pro- 
duces the  same  transformation. 

3.  The  biliary  acids,  the  taurocholic  and  glycocholic, 
are  susceptible,  as  we  know  by  the  works  of  Strecker,  of 
being  resolved  by  hydratation  into  taurine  and  cholalic 
acid,  or  into  glycocoll  and  cholalic  acid.  All  that  need 
be  done,  in  order  to  attain  this  result,  is  to  boil  it  for  a 
sufficient  time  with  baryta.  The  same  effect  of  decom- 
position is  observed  when  bile  is  left  to  itself,  to  undergo 
spontaneous  change.  Taurocholic  acid  is  transformed 
more  easily  than  its  accompanying  acid.  This  easy 
transformation,  due  to  the  intervention  of  ferments,  has 
for  a  long  time  obscured  the  chemical  history  of  bile. 

4.  The  decomposition  of  hippuric  acid,  in  the  urine  of 
the  herbivora,  into  benzoic  acid  and  glycocoll  is  a  reac- 
tion of  the  same  order. 

5.  This  is  the  case  also  with  two  vegetable  glucosides, 
phiilyrin,  contained  in  the  bark  of  the  Phillyrea  latifolia^ 
and  popitlin,  from  the  bark  of  the  aspen.  These  sub- 
stances are  attacked  neither  by  beer-yeast  nor  by 
emulsin  ;  but  when  they  are  placed  under  the  condi- 
tions of  lactic  fermentation,  they  are  resolved,  the  one 
into  glucose  and  phillygenin — 

U 


3o6 


ON  FERMENTATION. 


Q27  QU   ^    YL'i   Q   ^   Q6   IIVZ   QG   ^    Q21  ^24 

Phillyrin.  Glucose.  Phillygenin. 

the  other  into  glucose,  sahgenin,  and  benzoic  acid — 

Ci^        (cr  H»  O)  O^  +       O   =  O^  +  O  H«  O^ 

Populin  or  benzoyl  salicin.  Salicin.  Benzoic  acid. 

The  saHcin  formed  at  first  is  resolved  in  its  turn  into 
glucose  and  saligenin. 

6.  Infusions  of  gall-nuts,  left  to  themselves,  ferment ; 
the  tannin  disappears,  and  is  replaced  by  gallic  and 
ellagic  acid,  and  glucose. 

Strecker,  considering  the  tannin  as  a  giucoside,  repre- 
sented its  decomposition  by  the  equation — 

or  H-2  0'7  +  4  H2  O  =  0«  +  3  C7  H«  O^ 

Tannin.  Glucose.  Gallic  acid. 

7.  According  to  M.  Fremy,  pectose  is  accompanied,  in 
the  vegetable  tissues  in  which  it  is  found,  by  a  ferment, 
pectose,  sometimes  soluble  and  at  others  insoluble,  which 
possesses  the  property  of  transforming  pectose  and 
pectin  into  pectic  and  metapectic  acids  successively. 

Pcctic  fermentation  plays  an  important  part  in  the 
conversion  of  ripe  fruits  into  an  over-ripe,  half-rotten, 
or  sleepy"  state.  It  also  assists  in  the  formation 
of  vegetable  jellies.  In  fact,  the  transformation  of 
the  natural  juices  of  fruits  into  jellies  is  a  result  of 
the  metamorphosis  of  pectin,  contained  in  these  juices, 
into  the  pectosic  and  pectic  acids.  If,  then,  a  natural 
juice — that  of  the  currant,  for  example — does  not  con- 
tain pectose,  we  must  add  to  it  another  juice,  or  pulp, 


SOLUBLE  FERMENTS. 


which  contains  it,  in  a  soluble  or  unsoluble  form, 
remembering  that  boiling,  by  coagulating  the  ferment, 
renders  it  inactive.  Pectic  fermentation  is  effected  at 
about  35°  C.  (95°  R).  (Fremy,  Ann.  Chim.  Phys.  (3),  vol. 
24,  p.  I.) 


308 


ON  FERMENTATION. 


CHAPTER  III. 

ON  THE  ORIGIN  OF  FERMENTS. 

The  question  of  the  origin  of  ferments  is  intimately 
connected  with  that  of  spontaneous  generation.  In  fact, 
from  the  time  of  Van  Helmont  and  others,  who,  even 
in  the  seventeenth  century,  gave  directions  for  the  produc- 
tion of  mice,  frogs,  eels,  &c.,  the  partisans  of  this  mode 
of  generation  have,  by  the  progress  of  the  tendency  to 
examine  into  the  causes  of  things,  been  driven  from  the 
larger  animals  or  plants  visible  to  the  naked  eye,  to  the 
smallest  living  productions,  which  we  can  observe  only 
by  the  aid  of  the  microscope.  But  ferments  are  found 
among  these  inferior  microscopic  organisms.  Redi,  a 
member  of  the  Academy  of  Cimento,  showed  that  the 
worms  in  putrefied  flesh,  which  were  at  first  thought  to 
be  of  spontaneous  origin,  are  only  the  larvse  from  the 
eggs  of  flies,  and  that  all  that  was  necessary  to  prevent 
entirely  the  birth  of  these  larvae,  was  to  surround  the 
decomposing  meat  with  fine  gauze ;  he  was  the  first 
to  ascertain  that  parasitic  animals  are  sexual  and  able 
to  lay  eggs. 

The  invention  of  the  microscope,  and  the  numerous 
observations  by  which  it  was  followed,  towards  the  end 
of  the  seventeenth,  and  the  commencement  of  the  eight- 
eenth, century,  gave  fresh  impulse  to  the  doctrine  of 


ON  THE  ORIGIN  OF  FERMENTS. 


309 


spontaneous  generation,  which  had  lost  all  credit  in  ques- 
tions concerning  the  origin  of  living  beings  of  a  higher 
order. 

The  question  now  was  how  to  explain  the  origin  of 
the  various  living  productions,  revealed  by  the  micro- 
scope in  infusions  of  vegetable  and  animal  substances, 
among  which  no  apparent  symptom  of  sexual  genera- 
tion could  then  be  found. 

The  subject  was  studied  for  the  first  time  in  a  scien- 
tific manner  by  Needham,  who  published  in  1745,  in 
London,  a  work  on  this  subject.  This  observer  did  for 
infusoria  what  had  already  been  done  for  the  higher 
organisms.  He  protected,  or  rather  endeavoured  to  pro- 
tect, vegetable  or  animal  infusions  from  the  action  of 
germs,  seeds,  or  any  other  agents  of  mutiplication  which 
could  come  from  without.  At  the  same  time  he  destroyed 
by  a  physical  agent,  heat,  the  germs  which  might  be 
supposed  to  exist  beforehand  in  the  liquid.  Under  these 
conditions,  either  living  beings  will  be  produced  in  the 
midst  of  the  infusion,  or  none  will,  be  found  there  ;  in 
the  former  case,  it  must  be  admitted  that  these  or- 
ganisms are  developed  in  the  medium  which  is  suitable 
to  them,  without  the  intervention  of  any  germ  ;  in  the 
second,  that  the  doctrine  of  spontaneous  generation  is 
false.  In  reality,  the  question  can  only  be  resolved  in 
this  manner,  and  all  experimenters  who  have  entered 
upon  it  from  Needham's  time  to  the  present  day  ought 
to  have  made  use  of  it. 

The  serious  and  grave  difficulty,  on  which,  during  this 
period,  all  discussions  raised  between  heterogenists  and 
panspermists  have  turned,  is  so  to  arrange  the  experi- 
ments as  to  remove  every  suspicion  of  the  intervention 


310 


ON  FERMENTATION. 


of  germs  brought  from  without,  or  pre-existing  in  the 
Hquid. 

If  the  result  is  negative,  if  when  all  precautions  that 
seem  to  be  necessary  have  been  taken,  and  all  causes 
of  error  have  been  removed,  there  is  no  formation  of 
infusoria,  it  will  be  difficult  to  raise  any  serious  objection 
to  the  inevitable  conclusion,  provided  that  the  methods 
employed  for  the  purpose  of  eliminating  the  pre-existing 
germs  are  not  of  such  a  nature  as  to  modify  the  medium, 
and  to  render  it  unfit  for  the  development  and  the  nutri- 
tion of  living  organisms.  If,  on  the  contrary,  we  still 
meet  with  the  birth  of  living  beings,  the  suspicion  will 
always  revive  that  the  experiment  has  been  badly  per- 
formed, and  that  a  contrary  result  would  have  been 
obtained  by  conducting  it  more  carefully.  The  hetero- 
genists,  therefore,  find  themselves  in  a  more  disadvan- 
tageous situation  than  their  opponents,  and  notwith- 
standing the  success  which  they  may  obtain,  they  will 
never  convince  them. 

We  think  therefore  that  it  is  useless  to  give  here  a 
detailed  account  of  their  minute  researches  ;  they  must 
be  consulted  in  the  original  memoirs.  A  single  experi- 
ment which  proves y  by  a  negative  result ^  that  organic 
infusions^  protected  from  germs  from  zvithont,  do  not  give 
birth  to  inftcsoria,  is  worth  more,  scientifically  speaki7tg^ 
than  ten  experiments  tending  to  establish  the  contrary 
opinion. 

If,  therefore,  we  pass  over  the  details  of  the  funda- 
mental experiments  of  the  heterogenists,  and  speak  of 
those  the  results  of  which  are  conformable  to  the  ideas 
of  the  panspermists,  it  will  not  be  in  a  spirit  of  par- 
tiality.   We  are  convinced  that  the  latter  are  the  only 


ON  THE  ORIGIN  OF  FERMENTS.  3 II 

ones  free  from  all  objections,  the  relative  skill  of  the 
operators  being  disregarded,  and  considered  as  nothing 
in  the  estimate  formed.  We  may,  however,  say,  that 
M.  Pasteur's  researches  may  serve  as  a  model  for  all 
those  who  may  wish  to  conduct  investigations  of  this 
kind,  whatever  may  be  the  preconceived  opinion  by 
which  they  are  guided.  By  their  precision,  and  the  care 
taken  to  remove  every  source  of  error,  they  leave  nothing 
to  be  desired. 

As  the  results  obtained  by  M.  Pasteur  lead  him  to 
deny  spontaneous  generation,  his  opponents  must  above 
all  prove  that  he  is  mistaken,  by  adopting  the  same 
rigorous  experimental  conditions. 

Needham's  experiments,  of  which  we  have  before 
spoken,  which  led  this  observer  to  admit  and  sustain  the 
doctrine  of  spontaneous  generation,  consisted  essentially 
in  placing  organic  substances  which  were  capable  of 
decomposition,  in  vessels  hermetically  sealed,  which 
were  subsequently  submitted  to  a  high  temperature,  in 
order  to  destroy  the  pre-existing  germs. 

The  work  of  the  English  writer  attracted  great  notice 
on  account  of  the  support  of  Buffon,  whose  ideas  he 
upheld. 

Soon  after  began  the  great  controversy  between  Need- 
ham  and  Spallanzani,  a  celebrated  Italian  physiologist. 
The  latter,  in  his  treatises  on  animal  and  vegetable 
physics,  translated  by  Sennebier,  in  1777,  refuted  by 
experiment  the  conclusions  arrived  at  by  Needham. 

The  controversy  turned  principally  on  this  point ; 
Spallanzani  was  not  satisfied  with  heating  the  hermeti- 
cally sealed  vessels  containing  the  infusions,  for  several 
minutes,  merely  the  time  zvhieh  is  required -io  cook  a  heiis 


ON  FERMENTATION, 


egg,  and  io  destroy  the  germs,  as  Needham  expresses  it ; 
but  he  kept  them  for  the  space  of  an  hour  in  boiHng 
water.  He  then  observed  no  production  of  infusoria. 
But,  objects  the  EngHsh  observer,  from  the  manner  in 
which  he  treated  and  put  to  the  torture  his  nineteen 
vegetable  infusions,  it  is  evident  that  he  not  only  much 
weakened,  or  perhaps  totally  destroyed,  the  vegetative 
force  of  the  substances  infused,  but  also  entirely  cor- 
rupted, by  the  exhalations  and  the  odour  of  the  fire,  the 
small  portion  of  air  which  remained  in  the  empty  part 
of  his  vessels.  It  is  not  therefore  surprising  that  his 
infusions,  thus  treated,  gave  no  signs  of  life.  Such  must 
necessarily  have  been  the  case. 

This  idea,  that  the  action  of  the  temperature  of  boil- 
ing water  destroys  the  vegetative  force  of  infusions,  is 
maintained  even  at  the  present  day,  and  has  served  as 
an  argument  to  the  heterogenists ;  as  they  were  unable 
to  attack  the  material  correctness  of  Pasteur's  experi- 
ments, they  did  not  accept  the  conclusions  which  he 
sought  to  derive  from  them. 

We  find  also  in  the  passage  just  cited,  the  necessity 
for  the  experiments  made  by  Schwann  and  Helmholtz 
on  calcined  air,  and  for  those  of  Schroeder  and  F.  Dusch, 
on  strained  air. 

The  objection  of  a  possible  change  in  the  air  contained 
in  the  phial,  under  the  influence  of  prolonged  boiling, 
in  presence  of  organic  substances,  was  a  serious  one  at 
the  time  that  it  was  brought  forward  ;  it  becomes  more 
so,  when  we  know  that  the  air  confined  over  preserved 
meats,  prepared  by  Appert's  process,  contains  no  oxygen. 
It  was,  therefore,  absolutely  necessary  to  place  the  infu- 
sions in  contact  with  air  in  a  normal  condition,  after 


ON  THE  ORIGIN  OF  FERMENTS. 


that  boiling  had  deprived  them  of  their  pre-existing 
germs,  avoiding  at  the  same  time  any  new  germs  brought 
by  the  air. 

For  this  purpose,  Dr.  Schwann  heated  flasks  con- 
taining the  infusions,  until  the  destruction  of  the  germs 
was  insured  ;  but  his  flask  was  not  closed  ;  it  commu- 
nicated freely  with  the  surrounding  air  by  means  of  a 
glass  tube  bent  in  the  form  of  an  U,  and  heated,  in  one 
part  of  its  length,  by  means  of  a  bath  of  fusible  alloy. 
Under  these  conditions,  the  air  may  be  renewed  in  the 
flasks,  but  the  fresh  atmospheric  air  admitted  has  under- 
gone, like  the  infusion,  the  action  of  heat,  which  destroys 
the  germs. 

Schwann's  experiment  was  very  decisive,  as  to  broth 
made  from  meat ;  and  the  negative  result  (no  develop- 
ment of  infusoria)  was  quite  satisfactory.  But  it  was 
not  the  same  with  analogous  trials  on  alcoholic  fermenta- 
tion, which  gave  contradictory  results. 

Ure  and  Helmholtz  repeated  and  multiplied  these 
experiments  with  the  same  success. 

To  obviate  the  objection  of  a  possible  change  by  heat, 
in  a  mysterious  and  undefined  principle,  different  from 
germs,  but  whose  presence  in  the  air  was  necessary  to 
the  production  of  infusoria,  Schultz  (Ann.  des  Sciences 
Naturelles,  vol.  8,  (2),  1837)  caused  the  renewed  air  to 
pass  through  energetic  chemical  re-agents,  such  as  con- 
centrated sulphuric  acid.  He  half  filled  a  glass  vessel 
with  distilled  water  containing  various  animal  and  vege- 
table substances  ;  then  stopped  the  vessel  with  a  cork 
through  which  passed  two  bent  tubes,  and  exposed  the 
apparatus  thus  arranged  to  the  temperature  of  boiling 
water.   Then,  while  the  vapour  was  still  escaping  through 


i 


3^4 


ON  FERMENTATION. 


the  tubes  of  which  we  have  just  spoken,  he  adapted 
to  each  of  them  a  Liebig's  bulb  apparatus,  , one  contain- 
ing concentrated  sulphuric  acid,  and  the  other  concen- 
trated caustic  •  potash.  The  high  temperature  must 
necessarily  have  destroyed  every  living  thing,  all  the 
germs  that  might  happen  to  be  in  the  inside  of  the 
vessel,  or  of  its  appendages,  and  the  communication 
from  without  was  intercepted  by  the  sulphuric  acid  on  one 
side  and  the  potassa  on  the  other.  Nevertheless,  it  was 
easy  to  renew  by  aspiration  at  the  end  of  the  apparatus 
which  contained  the  potassa,  the  air  thus  enclosed,  and 
the  fresh  quantities  of  this  fluid  which  were  introduced 
could  not  carry  with  them  any  living  germ,  for  they  were 
forced  to  pass  through  a  bath  of  concentrated  sulphuric 
acid.  M.  Schultze  placed  the  apparatus  thus  arranged 
at  a  well  lighted  window,  side  by  side  with  an  open 
vessel,  which  contained  an  infusion  of  the  same  organic 
substances ;  then  he  was  careful  to  renew  the  air  in  his 
apparatus  several  times  a  day  for  more  than  two  months, 
and  to  examine  with  the  microscope  what  took  place  in 
the  infusion.  The  open  vessel  was  soon  found  filled 
with  vibrios  and  monads,  to  which  were  soon  added 
polygastric  infusoria  of  a  larger  size,  and  even  rotifers ; 
but  by  the  most  attentive  observation  he  could  not  dis- 
cover the  least  trace  of  infusoria,  confervas,  or  mildews, 
in  the  infusion  contained  in  the  apparatus. 

The  latest  researches  of  Schroeder  and  V.  Dusch 
(1854-1859)  tended  to  raise  another  objection,  the  pos- 
sible change  in  a  special  principle  in  the  air,  by  a  re-agent 
as  energetic  as  sulphuric  acid.  Guided  by  the  experi- 
ments of  Loewel,  who  ascertained  that  common  air, 
when  it  had  been  previously  filtered  through  cotton,  was 


ON  THE  ORIGIN  OF  FERMENTS. 


unfit  to  cause  the  crystallization  of  supersaturated  solu- 
tions of  sodium  sulphate,  they  placed  one  of  the  tubes 
of  Schutze's  apparatus  in  communication  with  a  tube 
3  centimetres  (i'i8  in.)  in  diameter,  and  from  50  to  60 
centimetres  (19*68  to  23*62  in.)  in  length,  filled  with 
cotton-wool.  The  other  tube  was  connected  with  an 
aspirator. 

When  the  liquid,  the  interior  of  the  flask,  and  the 
tubes,  had  been  been  deprived  of  air  by  boiling,  the 
apparatus  was  removed  to  its  place,  and  the  aspiration 
continued  night  and  day. 

The  two  observers  thus  proved  that  meat  to  which 
water  had  been  added,  the  wort  of  beer,  urine,  starch, 
paste,  and  the  various  materials  of  milk  taken  sepa- 
rately, remained  intact  in  the  filtered  air. 

On  the  contrary,  milk,  meat  without  water,  and  the 
yolk  of  egg,  grew  putrid  as  rapidly  as  in  common  air. 

The  result  of  these  experiments  is,  that  there  are 
spontaneous  decompositions  of  organic  substances  which 
require  nothing  but  the  presence  of  oxygen  gas  to  cause 
them  to  commence  ;  while  others  need,  besides  oxygen, 
the  presence  in  the  atmospheric  air  of  those  tmknoivn 
thingSy  which  are  destroyed  by  heat  or  sulphuric  acid, 
or  are  retained  by  the  cotton. 

The  two  observers  do  not  then  decide  on  the  nature  of 
these  things,  and  do  not  assert  categorically  that  they 
are  germs,  and,  in  reality,  nothing  allows  us  to  draw 
these  conclusions. 

M.  Pasteur's  experiments  (Memoire  on  the  Organic 
Corpuscles  which  exist  in  the  Atmosphere,  Ann. 
Chim.  Phys.  (3),  vol.  64,  p.  27)  have  advanced  the  ques- 
tion another  step,  by  proving  that  they  are  really  germs 


ON  FERMENTATION. 


of  ferments  and  infusoria,  which  are  destroyed  by  heat,  or 
arrested  by  the  sulphuric  acid  or  cotton  in  the  experi- 
ments alluded  to  above. 

M.  Pasteur  made  a  hole  in  a  window-shutter,  several 
metres  above  the  ground,  and  through  this  he  passed  a 
glass  tube  half  a  centemetre  {'196  in.)  in  diameter,  and 
containing  a  plug  of  soluble  cotton  i  centimetre 
(•39  in.)  in  length,  kept  in  its  place  by  a  spiral  platinum 
wire.  One  of  the  ends  of  this  tube  passed  into  the 
street  ;  the  other  communicated  with  a  continuous 
aspirator.  When  the  air  had  passed  for  a  sufficient 
time,  the  plug  of  cotton,  more  or  less  soiled  by  the  dust 
which  it  had  intercepted,  was  placed  in  a  small  tube  with 
the  mixture  of  alcohol  and  ether,  which  dissolves  gun- 
cotton.  It  was  left  for  the  space  of  a  day.  All  the 
dust  was  deposited  at  the  bottom  of  the  tube,  where  it 
is  easy  to  wash  it  by  decantation,  without  any  loss,  if 
care  is  taken  to  separate  each  washing  by  an  interval  of 
repose  of  from  twelve  to  twenty  hours.  The  deposit, 
and  the  liquid  which  covers  it,  are  put  in  a  watch-glass 
together ;  after  the  evaporation  of  the  alcohol,  the 
remainder  is  placed  in  water,  and  examined  with  the 
microscope.  M.  Pasteur  also  made  use  of  ordinary 
sulphuric  acid  in  order  to  moisten  the  ddst.  This  agent 
had  the  effect  of  separating  the  grains  of  starch  and 
calcium  carbonate,  which  are  always  found  in  greater 
or  less  quantities  in  deposits  collected  on  the  plug  of 
cotton. 

Figs.  24  and  25  represent  organic  corpuscules,  asso- 
ciated will  amorphous  particles,  as  seen  through  the 
microscope,  under  a  power  of  350  diametres  ;  the  liquid 
containing  tliem  was  common  sulphuric  acid. 


ON  THE  ORIGIN  OF  FERMENTS.  317 

Fig.  24  applies  to  dust  collected  from  the  25th  to  the 
26th  of  June,  i860;  fig.  25  to  dust  from  the  very  intense 
fog  of  January,  1861. 

It  was  not  enough  to  discover  with  the  microscope 
organic  particles  mixed  with  amorphous  substances,  but 


FiOs.  24  and  25. — Organic  corpuscules  of  dust,  mixed  with  amorphous  particles. 

it  was  necessary  to  prove  that  these  particles  really  con- 
sisted of  fertile  germs,  capable  of  producing  the  infusoria 
which  are  developed  in  such  abundance  in  organic  liquids 
exposed  to  the  air. 

For  this  purpose,  M.  Pasteur  arranged  the  experiment 
in  the  following  manner  : — 

Into  a  flask  capable  of  containing  from  250  to  300 
cubic  centimetres  (15  to  18  cub.  in.),  he  introduced  100 
or  150  cubic  centimetres  (6  to  9  cub.  in.)  of  albuminous 
saccharine  water,  prepared  in  the  following  propor- 
tions : — 

Water  100 
Sugar  10 

Albuminoid  and  mineral  matter  from  beer-yeast  '2  to  7. 

The  neck  of  the  drawn-out  neck  flask  communicated 
with  a  platinum  tube,  as  shown  in  Fig.  26.  In  this 
first  stage  of  the  experiment  the  T-shaped  tube  with 


3i8 


ON  FERMENTATION. 


three  stop-cocks  is  removed,  and  its  place  supplied  by  a 
simple  india-rubber  connecting  piece.  The  platinum 
tube  is  raised  to  a  red  heat  by  means  of  a  small  gas 
furnace.  The  liquid  is  boiled  for  two  or  three  minutes, 
and  is  then  allowed  to  grow  completely  cold.  It  is 
filled  with  common  air,  at  the  ordinary  pressure  of  the 
atmosphere,  but  which  has  been  wholly  exposed  to  a 
red  heat;  then  the  neck  of  the  flask  is  hermetically  seajed. 

This,  being  thus  prepared,  and  detached,  •  is  placed 
in  a  stove  at  a  constant  temperature  of  about  30°  C. 
(86°  F.) ;  it  may  be  kept  there  for  any  length  of  time 
without  the  least  change  in  the  liquid  which  it 
contains.  It  preserves  its  limpidity,  its  smell,  and 
and  its  weak  acid  reaction  ;  even  a  very  slight  absorp- 
tion of  oxygen  is  mainly  to  be  observed.  M.  Pasteur 
affirms  that  he  never  had  a  single  experiment  which  was 
arranged  as  described  above,  which  yielded  a  doubtful 
result ;  while  water  of  yeast  mixed  with  sugar,  and 
boiled  for  two  or  three  minutes,  and  then  exposed  to 
the  air,  was  already  in  evident  process  of  decomposition 
in  a  day  or  two,  and  was  found  to  be  filled  with  bacteria 
and  vibrios,  or  covered  with  mucors.  These  experiments 
are  directly  opposed  to  those  of  Messrs.  Pousset,  Man- 
tcgazzo,  Joly,  and  Musset. 

It  is  therefore  clearly  proved  that  sweetened  yeast 
water,  a  liquid  very  liable  to  be  decomposed  by  the 
contact  of  common  air,  may  be  preserved  for  years 
unaltered  when  it  has  been  exposed  to  the  action  of 
calcined  air,  after  having  been  allowed  to  boil  for  a  few 
minutes  (two  or  three).* 

*  M.  Pasteur  has  pointed  out  a  cause  of  want  of  success,  which  has  led 
many  experimenters  into  error ;  by  showing  that  the  mercury  of  a  mercurial 


i 


320  ON  FERMENTATION. 

This  being  determined,  M.  Pasteur  adapted,  by  means 
of  an  india-rubber  tube,  the  closed  point  of  his  flask 
filled  with  sweetened  yeast  water,  which  had  been  kept 
for  two  or  three  months  in  a  heated  stove,  without  any 
development  of  organisms,  to  an  apparatus  arranged  like 
that  in  figure  26. 

The  pointed  end  of  the  flask  passed  into  a  strong  glass 
tube  10  or  12  miUimetres  ('39  to  '46  in.)  in  its  inner  dia- 
meter, within  which  he  had  placed  a  piece  of  tube  of  small 
diameter  open  at  both  ends,  free  to  slip  in  the  larger 
tube,  and  enclosing  a  portion  of  one  of  the  small  plugs 
of  cotton  loaded  with  dust.  The  larger  glass  tube  is 
bound  to  a  brass  tube  in  form  of  a  T,  furnished  with 
stop-cocks,  one  of  which  communicates  with  the  air- 
pump,  another  with  the  heated  platinum  tube,  and  the 
third  with  the  flask,  by  means  of  the  large  tube  which 
contains  the  smaller  one  with  the  cotton.  These  various 
parts  are  joined  together  by  means  of  india-rubber. 

The  experiment  is  commenced  by  exhausting  the  air, 
after  having  closed  the  stop-cock  connected  with  the 
red-hot  metallic  tube.  This  being  afterwards  opened, 
allows  calcined  air  to  enter  the  tubes  slowly ;  this  opera- 
tion (exhaustion  and  readmission  of  calcined  air)  is 
repeated  several  times.  The  point  of  the  flask  is  then 
broken  off  within  the  india-rubber,  and  the  small  tube 
containing  the  dust  is  allowed  to  slip  into  the  flask,  the 
neck  of  which  is  again  sealed  with  the  lamp.  As  an 
additional  proof,  and  to  obviate  all  objections,  the  same 
arrangements  were  made  with  similar  flasks,  prepared 

trough  is  a  complete  receptacle  for  living  organisms,  and  consequently  that  all 
experiments  made  with  such  a  trough  must  necessarily  induce  a  development 
of  infusoria. 


.i 


ON  THE  ORIGIN  OF  FERMENTS. 


321 


like  the  preceding,  but  with  this  difference,  that  instead 
of  cotton  charged  with  atmospheric  dust,  there  was 
substituted  a  small  piece  of  tube  containing  calcined 
asbestos ;  (as  an  additional  precaution,  it  had  been 
ascertained  that  calcined  asbestos,  loaded  with  atmos- 
pheric dust,  by  the  same  means  as  the  cotton,  gave 
identical  results). 

The  following  are  the  observations  obtained  constantly 
by  M.  Pasteur  :— 

In  all  the  flasks,  into  which  dust  collected  from  the 
air  were  introduced,  i.  Organic  productions  began  to 
make  their  appearance  in  the  liquid  after  24,  36,  or 
48  hours  at  the  most.  This  was  precisely  the  time 
necessary  for  the  same  phenomena  to  appear  in 
sweetened  yeast-water  exposed  to  contact  with  the 
atmosphere. 

2.  The  products  observed  are  of  the  same  kind  as 
those  which  are  seen  to  make  their  appearance  in  the 
liquid  when  left  freely  exposed  to  the  air,  such  as 
mucors,  common  mucidines,  torulacei,  bacteria,  and 
vibrios  of  the  smallest  species,  the  largest  of  which,  the 
monas  Ims,  is  only  '004  millimetre  (•000157  in.)  in 
diameter. 

It  was  a  remarkable  circumstance  that  M.  Pasteur 
never  saw  any  alcoholic  fermentation  appear,  although 
the  composition  of  the  liquid  employed  was  very  ap- 
propriate to  this  kind  of  change. 

When  the  water  of  yeast  is  replaced  by  urine,  the 
experiment  being  conducted  exactly  in  the  same  manner, 
we  always  notice  the  absence  of  any  change  as  long  as 
atmospheric  dust  has  not  been  introduced,  whilst  with 
the  addition  of  this,  numerous  organisms  are  developed, 


322 


ON  FERMENTATION. 


in  every  respect  similar  to  those  which  appear  and  are 
developed  in  urine  kept  in  the  open  air. 

If,  on  the  contrary,  the  experiment  be  repeated  with 
common  milk,  we  may  be  sure  that  it  will  in  every  case 
curdle,  and  become  putrid.  We  shall  observe  the  birth 
of  numerous  vibrios  of  the  same  species,  and  bacteria, 
and  the  oxygen  of  the  flask  will  disappear. 

M.  Pasteur  thinks  that  this  result,  so  different  from 
those  observed  in  other  liquids,  arises  only  from  the  fact 
that  milk  contains  germs  of  vibrios  which  resist  the 
boiling  heat  of  water.  To  prove  this,  he  boiled  milk, 
not  at  100°  C.  (212''  F.),  or  at  the  usual  pressure  of  the 
atmosphere,  but  at  110°  C.  (230''  F.),  under  a  greater 
pressure,  and  he  found  that  the  flasks  thus  prepared, 
and  hermetically  sealed,  could  be  kept  for  an  indefinite 
time  in  the  stove,  without  giving  rise  to  the  smallest 
production  of  mould  or  infusoria.  The  milk  preserves 
its  taste,  its  smell,  and  all  its  properties  ;  and  the  atmos- 
phere of  the  flask  is  only  slightly  modified  in  its  com- 
position. (After  forty  days  there  were  still  found  18*37 
volumes  of  oxygen  per  100  parts  of  air.) 

This  difference  between  milk  and  urine,  or  sweetened 
yeast-water,  must  be  attributed  to  the  alkaline  condition 
of  the  former  medium,  whereas  the  two  others  are 
acid. 

In  fact,  if  we  previously  neutralize  the  acid  of  the 
sweetened  yeast-water,  by  means  of  calcium  carbonate, 
we  obtain  organisms  under  the  same  conditions  of  the 
experiment  as  those  under  which  they  were  not  before 
developed. 

These  facts  led  M.  Pasteur  to  make  researches  on  the 
comparative  action  of  temperature  on  the  fecundity  of 


I 


324 


ON  FERMENTATION. 


the  spores  of  the  mucidines,  and  of  the  germs  which 
exist  suspended  in  the  atmosphere. 

The  following  is,  in  few  words,  the  method  followed 
by  him.  He  passed  a  small  portion  of  asbestos  over 
the  small  heads  of  the  moulds  which  he  wished  to  study  ; 
he  then  placed  this  asbestos,  covered  with  spores,  in  a 
small  glass  tube,  which  he  introduced  into  an  U  tube 
(Fig.  27)  of  larger  diameter,  in  which  the  smaller  tube 
could  move  freely ;  one  of  the  extremities  of  the  U  tube 
is  joined  by  india-rubber  to  a  metal  tube  in  form  of  a 
T,  with  stop-cocks.  One  of  these  cocks  communicated 
with  the  air-pump,  another  with  a  red-hot  platinum  tube. 
The  other  extremity  has  an  india-rubber  tube  which  is 
connected  with  the  flask  into  which  the  spores  are  to  be 
introduced  ;  this  flask  is  hermetically  sealed,  and  has 
been  filled  with  calcined  air,  and  suitable  nutritious 
liquid  previously  raised  to  the  boiling  point.  Finally, 
the  U  tube  dips  into  a  bath  of  oil,  of  common  water,  or 
salt  water,  according  to  the  temperature  which  we  wish 
to  attain.  Between  the  U  tube  and  that  of  platinum, 
there  is  a  drying  tube  with  sulphuric  pumice-stone. 
When  all  the  apparatus  which  precedes  the  platinum 
tube  has  been  filled  with  calcined  air,  and  the  spores 
have  been  maintained  at  the  desired  temperature  for  a 
sufficient  time,  which  may  be  varied  at  pleasure,  the  point 
of  the  flask  is  broken  with  a  blow  of  a  hammer,  without 
unfastening  the  india-rubber  connecting  pieces  which 
attach  the  flask  to  the  U  tube  ;  then  inclining  to  a  proper 
angle,  this  latter  tube,  when  removed  from  its  bath,  the 
asbestos  with  its  spores  is  slipped  into  the  flask.  The 
flask  is  then  hermetically  scaled,  and  is  carried  to  the 
stove  at  20""  or  30°  C.  (68°  to  86°  F.).    The  experiment 


ON  THE  ORIGIN  OF  FERMENTS.  325 

with  the  dust  from  the  air  is  also  made  in  the  same 
manner  with  asbestos. 

Without  any  humidity,  the  fecundity  of  the  spores  of 
P enecillium  glattctmi  is  preserved  up  to  120°  C.  (248°  F.) 
and  even  a  httle  above — 125°  C.  (257''  F.).  It  is  the  same 
with  the  spores  of  the  other  common  mucidines.  At 
130°  C.  (266)°  F.),  the  power  of  developing  or  multi- 
plying is  destroyed  in  all  of  them.  The  limits  are  the 
same  for  the  dust  from  the  air. 

In  all  these  careful  experiments,  the  most  scrupulous 
precautions  were  taken  to  prevent  the  access  of  the 
slightest  portion  of  common  air.  But,  say  the  partisans 
of  heterogenesis,  if  the  smallest  portion  of  common 
air  develops  organisms  in  any  infusion  whatever,  it  must 
necessarily  be  the  case  that,  if  these  organisms  are  not 
spontaneously  generated,  there  must  be  germs  of  a 
multitude  of  various  productions  in  this  portion  of 
common  air,  however  small  it  may  be  ;  and  if  things 
were  so,  the  ordinary  air  would  be  loaded  with  organic 
matter  which  would  form  a  thick  mist  in  it. 

M.  Pasteur  has  shown  that  there  is  a  great  deal  of 
exaggeration  in  the  generally  received  opinion  that  even 
the  smallest  quantity  of  air  is  sufficient  to  develop 
multitudes  of  organisms  ;  that,  on  the  contrary,  there 
is  not  in  the  atmosphere  a  continuous  cause  of  these 
so-called  spontaneous  generations  ;  that  it  is  always 
possible  to  procure,  in  any  determined  place,  a  sufficient 
but  still  limited  quantity  of  common  air,  having  under- 
gone no  kind  of  modification,  whether  physical  or 
chemical,  and  nevertheless  quite  unsuited  to  set  up  any 
decomposing  action  in  a  liquid  eminently  putrescible. 
The  method  of  experimenting  is  very  simple.    Into  a 


326 


ON  FERMENTATION. 


flask  of  250  or  300  cubic  centimetres  (15  to  18  cub.  in.), 
150  cubic  centimetres  (9  cub.  in.)  of  a  liquid  that  has  a 
tendency  to  decomposition  are  introduced  ;  the  neck  of 
the  flask  is  drawn  out  with  the  lamp,  leaving  the  point 
open  ;  then  the  liquid  is  boiled  till  the  vapour  escaping 
from  the  extremity  has  expelled  all  the  air  ;  at  this 
moment  the  point  of  the  flask  is  closed  by  the  lamp,  by 
means  of  a  blow-pipe,  and  it  is  allowed  to  grow  cool.  The 
flask  then  contains  no  air ;  if  we  break  off  the  point  in 
any  particular  place,  the  air  re-enters  suddenly,  carrying 
into  it  the  germs  held  in  suspension  ;  it  is  again  closed 
with  the  lamp,  and  kept  in  a  stove  at  a  temperature  of 
20°  or  30°  C.  (68°  to  86°  R).  In  the  generality  of  cases, 
organisms  are  developed ;  these  organisms  are  even 
more  varied  than  if  the  liquid  were  freely  exposed  to 
the  air,  which  M.  Pasteur  explains  by  saying  that,  in  this 
case,  the  germs  in  small  number,  in  a  limited  volume  of 
air,  are  not  hindered  in  their  development  by  germs  in 
greater  number  or  more  precocious  in  their  fecundity, 
which  are  able  to  occupy  the  space,  and  leave  no  room  for 
them.  But  it  is  especially  important  to  notice  in  the  results 
obtained  by  this  method,  what  frequently  happens  many 
times  in  each  series  of  trials,  that  the  liquid  continues 
absolutely  intact,  however  long  it  may  have  remained 
in  the  stove,  as  if  it  had  been  filled  with  calcined  air. 
This  phenomenon  is  the  more  striking,  and  shows  it- 
self in  more  marked  proportions,  when  the  air  received 
into  the  flasks  is  taken  from  a  greater  height.^  Thus, 
among  twenty  flasks  opened  in  the  country,  eight  con- 
tained organic  productions ;  out  of  twenty  opened  on 
the  Jura,  only  five  contained  any  ;  and  out  of  twenty 
flasks  opened  at  Montanvcrt,  in  a  rather  high  wind, 


i 
1 


ON  THE  ORIGIN  OF  FERMENTS.  327 


blowing  from  the  deepest  gorges  of  the  "Glacier  des 
Bois,"  one  only  was  affected  by  any  change. 

We  may  also  draw  other  conclusions  from  this  series 
of  observations.  Since  the  putrescible  liquid,  which 
had  been  previously  boiled,  and  which  was  contained 


Fig.  28. — M.  Pasteur's  flask  to  deprive  the  air  of  its  germs. 

in  the  flasks,  was  filled  with  organic  productions  in  a 
great  number  of  instances,  after  the  introduction  of  a 
limited  quantity  of  air,  the  genetic  power  of  the  in- 
fusions had  not  been  destroyed  by  the  material  condi- 
tions of  the  experiments.  Besides,  this  objection,  which 
has  been  raised  ever  since  the  earliest  controversies 
between  the  heterogenists  and  the  panspermists,  has 
been  definitely  answered  by  an  experiment  made  by 
M.  Pasteur ;  he  received  in  a  flask,  exhausted  and 
deprived  of  living  germs  by  the  momentary  applica- 
tion of  a  sufficiently  high  temperature,  some  blood  at 
the  instant  that  it  left  the  organism,  and  without  allow- 
ing this  liquid,  which  is  so  peculiarly  putrescible,  to  come 
in  contact  with  air.  By  permitting  air  deprived  of 
germs,  either  by  calcination  or  simple  filtration,  to  enter 
the  flask,  and  then  hermetically  sealing  it,  he  found  that 


328 


ON  FERMENTATION. 


the  blood  was  preserved  for  an  indefinite  period  intact, 
although  it  had  not  been  exposed  to  heat. 

M.  Pasteur  has  also  shown  that  air  may  be  deprived 
of  germs  by  its  passage  through  a  capillary  tube  bent 
upon  itself.  It  is,  therefore,  sufficient,  in  most  cases,  to 
draw  out  the  neck  of  the  flask  so  as  to  form  a  very  long 
narrow  tube,  which  is  bent  in  several  directions,  as,  for 
example,  in  Fig.  28.  When  the  air  originally  contained 
in  it  had  been  expelled,  and  the  pre-existing  germs 
killed  by  prolonged  boiling,  the  flask  is  allowed  to  cool 
slowly. 

In  closing  our  account  of  M.  Pasteur  s  interesting 
memoir,  in  which  heterogenesis  was  driven  to  its  last 
intrenchments,  we  must  add  that  this  learned  chemist 
endeavoured  to  deprive  his  adversaries  of  one  of  their 
principal  arguments.  Experiments  on  spontaneous  gen- 
eration have  always  been  conducted  with  vegetable  or 
animal  infusions  ;  it  was  supposed  by  Needham,  Buflbn, 
and  Pouchet  that  organisms  were  only  thus  produced  at 
the  moment  of  expiring  nature,  when  the  elements  of 
the  beings  on  which  they  are  developed  are  entering 
into  new  chemical  combinations,  and  are  passing  fully 
through  the  phenomena  of  fermentation  or  putrefaction. 

In  other  words,  albuminoid  matters  preserve  in  some 
degree  a  certain  reserve  of  vitality,  which  would  allow 
them  to  become  organic  by  contact  with  oxygen,  when 
the  conditions  of  temperature  and  humidity  are  favour- 
able. Starting  with  the  idea  that  albuminoid  substances 
are  only  aliments  for  the  germs  of  infusoria,  mucidines, 
or  ferments,  M.  Pasteur  has  proved  directly  that  organic 
substances  may  be  replaced  by  those  which  are  purely 
mineral  or  artificial,  or,  at  least,  by  substances  on  which 


ON  THE  ORIGIN  OF  FERMENTS.  329 

this  imaginary  vegetative  force  cannot  be  supposed  to 
have  any  influence. 

We  have  spoken  elsewhere,  at  some  length,  of  the 
experiments  made  by  M.  Pasteur  and  M.  Raulin  on  the 
nutrition  and  development  of  ferments  and  of  muce- 
dines  in  artificial  media  composed  of  pure  sugar  candy, 
ammonium  tartrate,  and  phosphates. 

The  reader  will  doubtless  remember  an  observation 
of  M.  Pasteur's,  which  we  mentioned  in  passing,  but 
without  dwelling  upon  it.  When  he  introduced  into 
sweetened  yeast-water  that  had  been  previously  boiled 
and  preserved  in  calcined  air  atmospheric  dust  collected 
at  different  times  and  in  various  places,  he  never  met 
with  alcoholic  fermentation.  Yet  the  liquid  employed  is 
one  of  those  which  are  most  suitable  for  the  development 
of  alcoholic  ferments.  The  fact  may.  be  explained  by 
admitting  that  the  air  does  not  contain  spores  or  germs 
of  saccharomyces  or  apiculated  ferment ;  but  then  how 
can  we  explain  the  prompt  and  constant  appearance 
of  alcoholic  fermentation  in  the  juice  of  grapes  or  of 
fruits  in  general  }  The  cause  of  this  apparent  contra- 
diction is  very  simple.  It  is  not  the  air  which  brings 
the  germs  of  the  alcoholic  ferments  which  propagate  and 
multiply  so  rapidly  in  the  must  of  grapes,  or,  if  it  brings 
any,  they  are  in  so  minute  a  quantity  that  they  would 
not  be  sufficient  to  set  up  fermentation  in  so  short  a 
time.  These  germs  are  found  on  the  very  surface  of  the 
fruit,  on  the  grapes  which  contain  the  saccharine  liquid, 
the  decomposition  of  which  they  excite  as  soon  as  they 
are  placed  in  contact  with  it  when  the  fruit  is  pressed. 

In  order  to  prove  this,  M.  Pasteur  prepared  a  series  of 
forty  flasks,  with  sinuous  necks  like  those  which  we  have 
15 


330 


ON  FERMENTATION. 


described  above  (Fig.  28),  with  this  difference  only, 
that  the  neck  of  the  flask,  which  is  drawn  out  to  a  fine 
"swan's  neck''  is  not  the  only  one.  Each  flask  has  another 
straight  neck  closed  by  an  india-rubber  tube  furnished 
with  a  glass  stopper.  Into  these  forty  flasks  he  intro- 
duced limpid  and  filtered  grape  juice,  which,  after  being 
boiled,  like  all  liquids  which  are  slightly  acid,  remains 
intact,  although  the  extremity  of  the  neck  is  open.  On 
the  other  hand,  he  washed  in  a  few  cubic  centimetres 
of  water  part  of  a  bunch  of  grapes.  Under  the  micro- 
scope, we  may  perceive  in  this  water  the  existence  of  a 
multitude  of  organic  corpuscules,  resembling  so  closely 
as  to  be  indistinguishable  from  them,  either  spores  of 
minute  fungi,  alcoholic  ferment,  or  the  Mycoderma  vini. 
This  having  been  done,  M.  Pasteur  sowed  nothing  in 
ten  of  the  forty  flasks  ;  in  ten  others,  he  placed,  by 
means  of  the  second  tube,  some  drops  of  the  liquid  in 
which  the  grapes  had  been  washed.  In  a  third  series 
of  ten  other  flasks,  he  placed  some  drops  of  the  same 
liquid,  previously  raised  to  the  boiling  point,  and  then 
cooled.  Into  the  ten  remaining  flasks,  he  introduced 
one  drop  of  the  juice  of  a  grape  taken  from  those  that 
were  not  crushed. 

The  first  series  gave  no  product,  the  juice  having 
remained  unaltered.  In  the  second  series,  there  appeared 
some  flakes  of  mycelium,  some  alcoholic  ferment,  and, 
afterwards,  Mycoderma  villi ;  at  the  end  of  forty-eight 
hours  the  ten  flasks  were  in  full  fermentation,  if  they 
had  been  experimented  upon  at  the  temperature  of  the 
air.  The  third  series  had  not  a  single  flask  affected  ; 
the  liquid  remained  limpid.  In  the  fgurth,  a  single  flask 
was  changed.    The  conclusion   which  may  be  drawn 


ON  THE  ORIGIN  OF  FERMENTS.  33 1 


from  these  facts  is  simple*  M.  Bechamp  had  already 
proved,  by  former  experhi[ients,  that  grapes  bear  on  their 
surface  all  that  is  necessary  to  cause  saccharine  water  to 
ferment,  even  when  protected  from  the  air. 

With  the  question  of  the  origin  of  ferments  and  of 
spontaneous  generation,  is  connected  another,  which  may 
be  discussed  independently,  whatever  may  be  the  origin 
of  ferments,  whether  spontaneous  or  no ;  namely,  whether 
a  fermenting  organism  can  transform  itself  into  a  dif- 
ferent ferment,  endowed  with  distinct  active  properties, 
when  the  conditions  of  its  development  are  modified. 

It  is  evident  that  this  question  may  be  seriously  dis- 
cussed ;  it  is  connected  with  the  general  development 
theory  which  has  been  applied  to  higher  organisms  ;  it 
is,  therefore,  still  more  applicable  to  the  simplest  organ- 
isms of  the  living  creation.  The  facts  observed  are  not 
entirely  opposed  to  the  idea  of  a  transformation  of  fer- 
ments into  each  other;  we  have  even  had  occasion  to 
mention  some  which  seem  favourable  to  it.  In  the 
meanwhile,  these  facts  are  still  too  few  in  number  to  give 
rise  to  important  inferences ;  some  of  them  are  even 
contested.  We  shall  therefore  merely  call  attention  to 
this  branch  of  the  study  of  inferior  organisms ;  it  has 
already  been  the  object  of  valuable  researches,  which 
for  several  reasons  deserve  to  be  continued. 

*  Pasteur,  Comp.-Rend.,  vol.  75,  p.  781. 


opinions  of  the  Press  on  the  ^'International  Scientific  Series, 


I. 

Tyndall's  Forms  of  Water. 

I  vol.,  i2mo.    Cloth.    Illustrated  Price,  $1.50. 

**  In  the  volume  now  published,  Professor  Tyndall  has  presented  a  noble  illustration 
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scientific  vision,  his  singular  command  of  the  appropriate  language  of  exposition,  and 
the  peculiar  vivacity  and  grace  with  which  he  unfolds  the  results  of  intricate  scientific 
research." — N.  Y.  Tribune. 

"  The  *  Forms  of  Water,'  by  Professor  Tyndall,  is  an  interesting  and  instructive 
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"  This  series  is  admirably  commenced  by  this  little  volume  from  the  pen  of  Prof. 
Tyndall.  A  perfect  master  of  his  subject,  he  presents  in  a  style  easy  and  attractive  his 
methods  of  investigation,  and  the  results  obtained,  and  gives  to  the  reader  a  clear  con- 
ception of  all  the  wondrous  transformations  to  which  water  is  subjected." — Churchman. 


II. 

Bagehot  s  Physics  and  Politics, 

I  vol.,  i2mo.    Price,  $1.50. 

**  If  the  '  International  Scientific  Series  '  proceeds  as  it  has  begun,  it  will  more  than 
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Professor  Tyndall,  was  a  model  of  lucid  and  attractive  scientific  exposition  ;  and  now 
we  have  a  second,  by  Mr.  Walter  Bagehot,  which  is  not  only  very  lucid  and  charmingj^ 
but  also  original  and  suggestive  in  the  highest  degree.  Nowhere  since  the  publicatioii 
of  Sir  Henry  Maine's  'Ancient  Law,'  have  we  seen  so  many  fruitful  thoughts  sug- 
gested in  the  course  of  a  couple  of  hundred  pages.  .  .  .  To  do  justice  to  Mr.  Bage- 
hot's  fertile  book,  would  require  a  long  article.  W^ith  the  best  of  intentions,  we  are 
conscious  of  having  given  but  a  sorry  account  of  it  in  these  brief  paragraphs.  But  wo 
hope  we  have  said  enough  to  commend  it  to  the  attention  of  the  thoughtful  reader."— 
Prof  John  Fiske,  in  the  Atlantic  Monthly. 

"Mr.  Bagehot's  style  is  clear  and  vigorous.  We  refrain  from  giving  a  fuller  ac- 
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worth  their  while  to  peruse  the  book  for  themselves ;  and  we  sincerely  hope  that  the 
forthcoming  parts  of  the  'International  Scientific  Series*  will  be  as  interesting."— 
A  theneeum. 

"  Mr.  Bagehot  discusses  an  immense  variety  of  topics  connected  with  the  progress 
of  societies  and  nations,  and  the  development  of  their  distinctive  peculiarities;  and  hii 
book  shows  an  abundance  of  ingenious  and  original  thought." — Alfred  Russeli 
Wallace,  in  N attire. 


D.  APPLETON  &  CO.,  Publishers,  549  &  551  Broadway,  N.  Y. 


opinions  of  the  Press  oit  the  "'International  Scientific  S cries J^^ 


III. 

Foods. 

By  Dr.  EDWARD  SMITH. 

I  vol.,  i2ino.    Cloth.    Illustrated  Price,  $1.75. 

In  making  up  The  International  Scientific  Series,  Dr.  Edward  Smith  was  se- 
lected as  the  ablest  man  in  England  to  treat  the  important  subject  of  Foods.  His  services 
were  secured  for  the  undertaking,  and  the  little  treatise  he  has  produced  shows  that  the 
choice  of  a  writer  on  this  subject  was  most  fortunate,  as  the  book  is  unquestionably  the 
clearest  and  best-digested  compend  of  the  Science  of  Foods  that  has  appeared  in  our 
language. 

"  The  book  contains  a  series  of  diagrams,  dispfeying  the  effects  of  sleep  and  meals 
on  pulsation  and  respiration,  and  of  various  kinds  of  food  on  respiration,  which,  as  the 
results  of  Dr.  Smith's  own  experiments,  possess  a  very  high  value.  We  have' not  far 
to  go  in  this  work  for  occasions  of  favorable  criticism  ;  they  occur  throughout,  but  are 
perhaps  most  apparent  in  those  parts  of  the  subject  with  which  Dr.  Smith's  name  is  es- 
pecially linked." — London  Exauiiner. 

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and  a  part  of  which  could  not  easily  be  obtained  from  other  sources.  Its  interest  is  de. 
cidedly  enhanced  for  students  who  demand  both  clearness  and  exactness  of  statement, 
by  the  profusion  of  well-executed  woodcuts,  diagrams,  and  tables,  which  accompany  th^ 
volume.  .  .  .  The  suggestions  of  the  author  on  the  use  of  tea  and  coffee,  and  of  the  va- 
rious forms  of  alcohol,  although  perhaps  not  strictly  of  a  novel  character,  are  highly  in- 
structive, and  form  an  interesting  portion  of  the  volume." — N,  Y.  Tribune. 


IV. 

Body  and  Mind. 

THE  THEORIES  OF  THEIR  RELATION. 
By  ALEXANDER  BAIN,  LL.  D. 
I  vol.,  i2mo.     Cloth  Price,  $1.50. 

Professor  Bain  is  the  author  of  two  well-known  standard  works  upon  the  Science 
•f  Mind — "The  Senses  and  the  Intellect,"  and  "The  Emotions  and  the  Will."  He  is 
one  of  the  highest  living  authorities  in  the  school  which  holds  that  there  can  be  no  sound 
or  valid  psychology  unless  the  mind  and  the  body  are  studied,  as  they  exist,  together. 

'*  It  contains  a  forcible  statement  of  the  connection  between  mind  and  body,  study- 
ing their  subtile  interworkings  by  the  light  of  the  most  recent  physiological  investiga- 
tions. The  summary  in  Chapter  V.,  of  the  investigations  of  Dr.  Lionel  Beale  of  the 
embodiment  of  the  intellectual  functions  in  the  cerebral  system,  will  be  found  the 
freshest  and  most  interesting  part  of  his  book.  Prof.  Bain's  own  theory  of  the  connec- 
tion between  the  mental  and  tne  bodily  part  in  man  is  stated  by  himself  to  be  as  follows : 
There  is  *  one  substance,  with  two  sets  of  properties,  two  sides,  the  physical  and  the 
mental — a  double-faced  unity.'  While,  in  the  strongest  manner,  asserting  the  union 
of  mind  with  brain,  he  yet  denies  'the  association  of  union  in  place^*  but  asserts  the 
union  of  close  succession  in  time,'  holding  that  'the  same  being  is,  by  alternate  fits,  un- 
der extended  and  under  unextcnded  consciousness."  ' — Christian  Register. 


D.  APPT.ETON  &  CO.,  Pii1)1ishcrs,  549  c'v:  551  Broadway,  N.  Y. 


opinions  of  the  Press  on  the     International  Scientific  Series.'^'* 


V. 

The  Study  of  Sociology. 

By  HERBERT  SPENCER. 
I  vol.,  i2mo.    Cloth  Price,  $1.50. 

"The  philosopher  whose  distinguished  name  gives  weight  and  influence  to  this  vol- 
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and  departments.  There  is  a  fascination  in  his  array  of  facts,  incidents,  and  opinions, 
which  draws  on  the  reader  to  ascertain  his  conclusions.  The  coolness  and  calmness  of 
his  treatment  of  acknowledged  difficulties  and  grave  objections  to  his  theories  win  for 
him  a  close  attention  and  sustained  effort,  on  the  part  of  the  reader,  to  comprehend,  fol- 
low, grasp,  and  appropriate  his  principles.  This  book,  independently  of  its  bearing 
upon  sociology,  is  valuable  as  lucidly  showing  what  those  essential  characteristics  are 
which  entitle  any  arrangement  and  connection  of  facts  and  deductions  to  be  called  a 
science. " — Episcopalian, 

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style.    It  is  a  fascinating  work,  as  well  as  one  of  deep  practical  thought." — Bost,  Post, 

'*  Herbert  Spencer  is  unquestionably  the  foremost  living  thinker  in  the  psychological 
and  sociological  fields,  and  this  volume  is  an  important  contribution  to  the  science  of 
which  it  treats.  ...  It  will  prove  more  popular  than  any  of  its  author's  other  creations, 
for  it  is  more  plainly  addressed  to  the  people  and  has  a  more  practical  and  less  specu- 
lative cast.  It  will  require  thought,  but  it  is  well  worth  tlunking  about," — Albany 
Evening  yournal. 


VI. 

The  New  Chemistry. 

By  JOSIAH  P.  COOKE,  Jr., 

Erving  Professor  of  Chemistry  and  Mineralogy  in  Harvard  University. 
I  vol.,  i2mo.    Cloth.  Price,  $2.00. 

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**  This  admirable  monograph,  by  the  distinguished  Erving  Professor  of  Chemistry 
in  Harvard  University,  is  the  first  American  contribution  to  *  The  International  Scien- 
tific Series,'  and  a  more  attractive  piece  of  work  in  the  way  of  popular  exposition  upon 
a  difficult  subject  has  not  appeared  in  a  long  time.  It  not  only  well  sustains  the  char- 
acter  of  the  volumes  with  which  it  is  associated,  but  its  reproduction  in  European  coun- 
tries will  be  an  honor  to  American  science." — Neiv  York  Tribune, 

"  All  the  chemists  in  the  country  will  enjoy  its  perusal,  and  many  will  seize  upon  it 
as  a  thing  longed  for.  For,  to  those  advanced  students  who  have  kept  well  abreast  of 
the  chemical  tide,  it  offers  a  calm  philosophy.  To  those  others,  youngest  of  the  class, 
who  have  emerged  from  the  schools  since  new  methods  have  prevailed,  it  presents  a 
generalization,  drawing  to  its  use  all  the  data,  the  relations  of  which  the  newly-fledged 
fact-seeker  may  but  dimly  perceive  without  its  aid.  ...  To  the  old  chemists.  Prof. 
Cooke's  treatise  is  like  a  message  from  beyond  the  mountain.  They  have  heard  0/ 
changes  in  the  science ;  the  clash  of  the  battle  of  old  and  new  theories  has  stirred  them 
from  afar.  The  tidings,  too,  had  come  that  the  old  had  given  way ;  and  little  more  than 
this  they  knew.  .  .  .  Prof  Cooke's*  New  Chemistry'  must  do  wide  service  in  bringing 
to  close  sight  the  little  known  and  the  longed  for.  .  .  .  As  a  philosophy  it  is  elemen* 
tary,  but,  a^  a  book  of  science,  ordinary  readers  will  find  it  sufficiently  advanced."-^' 
Uiica  Morning  Herald. 

D,  APPLETON  &  CO.,  Publishers,  549  &  551  Broadway,  N.  Y. 


opinions  of  the  Press  on  the  ^^International  Scientific  Series,^^ 


VII. 

The  Conservation  of  Energy. 

By  BALFOUR  STEWART,  LL.  D„  F.  R,  S. 

With  an  Appendix  treating  of  the  Vital  and  Mejital  Applications  of  the  Doctrine. 
I  vol,,  i2mo.    Cloth.    Price,  $1.50. 

**  The  author  has  succeeded  in  presenting  the  facts  in  a  clear  and  satisfactory  manner, 
using  simple  language  and  copious  illustration  in  the  presentation  of  facts  and  prin- 
ciples, confining  himself,  however,  to  the  physical  aspect  of  the  subject.  In  the  Ap- 
pendix the  operation  of  the  principles  in  the  spheres  of  life  and  mind  is  supplied  by 
the  essays  of  Professors  Le  Conte  and  Bain." — Ohio  Farmer. 

"  Prof  Stewart  is  one  of  the  best  known  teachers  in  Owens  College  in  Manchester. 

**The  volume  of  The  International  Scientific  Series  now  before  us  is  an  ex- 
cellent illustration  of  the  true  method  of  teaching,  and  will  well  compare  with  Prof. 
Tyndall's  charming  little  book  in  the  same  series  on  *  Forms  of  Water,"  with  illustra- 
tions enough  to  make  clear,  but  not  to  conceal  his  thoughts,  in  a  style  simple  and 
brief." — Christian  Register,  Bostoii. 

y  The  writer  has  wonderful  ability  to  compress  much  information  into  a  few  words. 
It  is  a  rich  treat  to  read  such  a  book  as  this,  when  there  is  so  much  beauty  and  force 
combined  with  such  simplicity. — Eastern  Press, 


VJII. 

Animal  Locomotion; 

Or,  WALKING,  SWIMMING,  AND  FLYING. 

With  a  Dissertation  on  Aerona?4iics. 

By  J.  BELL  PETTIGREW,  M.  D.,  F.  R.  S.,  F.  R.  S.  E., 
F.  R.C.  P.E. 

I  vol.,  i2mo  Price,  $1.75. 

This  work  is  more  than  a  contribution  to  the  stock  of  entertaining  knowledge, 
though,  if  it  only  pleased,  that  would  be  sufificient  excuse  for  its  publication.  But  Dr. 
Pettigrew  has  given  his  time  ta«these  investigations  with  the  ultimate  purpose  of  solv- 
ing the  difficult  problem  of  Aeronautics.  To  this  he  devotes  the  last  fifty  pages  of  his 
book.  Dr.  Pettigrew  is  confident  that  man  will  yet  conquer  the  domain  of  the  air."— > 
N.  V.  yournal  of  Co^mnerce. 

"  Most  persons  claim  to  know  how  to  walk,  but  few  could  explain  the  mechanical 
principles  involved  in  this  most  ordinary  transaction,  and  will  be  sjirprised  that  the 
movements  of  bipeds  and  quadrupeds,  the  darting  and  rushing  motion  of  fish,  and  the 
erratic  flight  of  the  denizens  of  the  air,  are  not  only  anologous,  but  can  be  reduced  to 
similar  formula.  The  work  is  profusely  illustrated,  and,  without  reference  to  the  theory 
it  is  designed  to  expound,  will  be  regarded  as  a  valuable  addition  to  natural  history.'' 
—Omaha  Republic. 


D.  APPLETON  &  CO.,  Publishers,  549  &  551  Broadway,  N.  ¥♦ 


opinions  of  the  Press  on  the  ^^International  Scientific  Series. 


IX. 

Responsibility  in  Mental  Disease. 

By  HENRY  MAUDSLEY,  M.  D., 

Fellow  of  the  Royal  College  of  Physicians ;  Professor  of  Medical  Jurisprudence 
in  University  College,  London. 

I  vol.,  l2mo.    Cloth.    .    .    Price,  $1.50. 

"  Having  lectured  in  a  medical  college  on  Mental  Disease,  this  book  has  been  a 
feast  to  us.  It  handles  a  great  subject  in  a  masterly  manner,  and,  in  our  judgment,  the 
positions  taken  by  the  author  are  correct  and  well  sustained." — Pastor  and  People. 

*'  The  author  is  at  home  in  his  subject,  and  presents  his  views  in  an  almost  singu- 
larly clear  and  satisfactory  manner.  .  .  .  The  volume  is  a  valuable  contribution  to  one 
of  the  most  difficult,  and  at  the  same  time  one  of  the  most  important  subjects  of  inves- 
tigation at  the  present  day." — N.  Y.  Observer, 

*'It  is  a  work  profound  and  searching,  and  abounds  in  \fisdiQm J"— Pittsburg  Com- 
mercial, 

**  Handles  the  important  topic  with  masterly  power,  and  its  suggestions  are  prac- 
tical and  of  great  value." — Providence  Press, 

X. 

The  Science  of  Law. 

By  SHELDON  AMOS,  M.  A., 

Professor  of  Jurisprudence  in  University  College,  London  ;  author  of  **  A  Systematic 
View  of  the  Science  of  Jurisprudence,"  *'  An  English  Code,  its  Difficulties 
and  the  Modes  of  overcoming  them,"  etc.,  etc. 

I  vol.,  i2mo.    Cloth  Price,  $1.75. 

**The  valuable  series  of  'International  Scientific*  works,  prepared  by  eminent  spe- 
cialists, with  the  intention  of  popularizing  information  in  their  several  branches  of 
knowledge,  has  received  a  good  accession  in  this  compact  and  thoughtful  volume.  It 
is  a  difficult  task  to  give  the  outlines  of  a  complete  theory  of  law  in  a  portable  volume, 
which  he  who  runs  may  read,  and  probably  Professor  Amos  himself  would  be  the  last 
to  claim  that  he  has  perfectly  succeeded  in  doing  this.  But  he  has  certainly  done  much 
to  clear  the  science  of  law  from  the  technical  obscurities  which  darken  it  to  minds  which 
have  had  no  legal  training,  and  to  make  clear  to  his  '  lay '  readers  in  how  true  and  high  a 
sense  it  can  assert  its  right  to  be  considered  a  science,  and  not  a  mere  practice." — Thi 
Christian  Register. 

"  The  works  of  Bentham  and  Austin  are  abstruse  and  philosophical,  and  Maine's 
require  hard  study  and  a  certain  amount  of  special  training.  The  writers  also  pursue 
different  lines  of  investigation,  and  can  only  be  regarded  as  comprehensive  in  the  de- 
partments they  confined  themselves  to.  It  was  left  to  Amos  to  gather  up  the  result 
and  present  the  science  in  its  fullness.  The  unquestionable  merits  of  this,  his  last  book, 
are,  that  it  contains  a  complete  treatment  of  a  subject  which  has  hitherto  been  handled 
by  specialists,  and  it  opens  up  that  subject  to  every  inquiring  mind.  .  .  .  To  do  justice 
to  *  The  Science  of  Law  *  would  require  a  longer  review  than  we  have  «pace  for.  We 
have  read  no  more  interesting  and  instructive  book  for  some  time.  Its  themes  concern 
every  one  who  renders  obedience  to  laws,  and  who  would  have  those  laws  the  best 
possible.  The  tide  of  legal  reform  which  set  in  fifty  years  ago  has  to  sweep  yet  highei 
if  the  flaws  in  our  jurisprudence  are  to  be  removed.  The  process  of  change  cannot  be 
better  guided  than  by  a  well-informed  public  mind,  and  Prof.  Amos  has  done  greal 
service  in  materially  helping  to  promote  this  end." — ^Buffalo  Courier. 

D.  APPLETON  &  CO.,  Publishers,  549  &  551  Broadway,  N.  Y- 


opinions  of  the  Press  on  the  ^^International  Scientific  Series. 


XI. 

Animal  Mechanism, 

A  Treatise  on  Terrestrial  and  Aerial  Locojnotion, 
By  E.  J.  MAREY, 
Professor  at  the  College  of  France,  and  Member  of  the  Academy  of  Medicine, 
With  117  Illustrations,  drawn  and  engraved  under  the  direction  of  the  author. 
I  voL,  i2mo.    Cloth  Price,  $1.75 

"  We  hope  that,  in  the  short  glance  which  we  have  taken  of  some  of  the  most  im- 
portant points  discussed  in  the  work  before  us,  we  have  succeeded  in  interesting  our 
readers  sufficiently  in  its  contents  to  make  them  cm-ious  to  learn  more  of  its  subject- 
matter.    We  cordially  recommend  it  to  their  attention. 

**  The  author  of  the  present  work,  it  is  well  known,  stands  at  the  head  of  those 
physiologists  who  have  investigated  the  mechanism  of  animal  dynamics — indeed,  we 
may  almost  say  that  he  has  made  the  subject  his  own.  By  the  originality  of  his  con- 
ceptions, the  ingenuity  of  his  constructions,  the  skill  of  his  analysis,  and  the  persever- 
ance of  his  investigations,  he  has  surpassed  all  others  in  the  power  of  unveiling  the 
complex  and  intricate  movements  of  animated  h^m^s.'* -—Popular  Science  Mofithly. 


XII. 

History  of  the  Conflict  between 
Religion  and  Science. 

By  JOHN  WILLIAM  DRAPER,  M.  D.,  LL.  D/, 
Author  of    The  Intellectual  Development  of  Europe.*' 
I  vol.,  i2mo.   Price,  $1.75. 

"This  little  '  History'  would  have  been  a  valuable  contribution  to  literature  at  any 
iime,  and  is,  in  fact,  an  admirable  text-book  upon  a  subject  that  is  at  present  engross- 
ing the  attention  of  a  large  number  of  the  most  serious-minded  people,  and  it  is  no 
small  compliment  to  the  sagacity  of  its  distinguished  author  that  he  has  so  well  gauged 
the  requirements  of  the  times,  and  so  adequately  met  them  by  the  preparation  of  this 
volume.  It  remains  to  be  added  that,  while  the  writer  has  flinched  from  no  responsi- 
bility in  his  statements,  and  has  written  with  entire  fidelity  to  the  demands  of  truth 
and  justice,  there  is  not  a  word  in  his  book  that  can  give  offense  to  candid  and  fair- 
minded  readers." — N.  Y.  Evening  Post. 

**  The  key-note  to  this  volume  is  found  in  the  antagonism  between  the  progressive 
tendencies  of  the  human  mind  and  the  pretensions  of  ecclesiastical  authority,  as  devel- 
oped in  the  history  of  modern  science.  No  previous  writer  has  treated  the  subject 
from  this  point  of  view,  and  the  present  monograph  will  be  found  to  possess  no  less 
originality  of  conception  than  vigor  of  reasoning  and  wealth  of  erudition.  .  .  .  The 
method  of  Dr.  Draper,  in  his  treatment  of  the  various  questions  that  come  up  for  dis- 
cussion, is  marked  by  singular  impartiality  as  well  as  consummate  ability.  Through- 
out his  work  he  maintains  the  position  of  an  historian,  not  of  an  advocate.  ^  His  tone  is 
tranquil  and  serene,  as  becomes  the  search  after  truth,  with  no  trace  of  the  impassioned 
ardor  of  controversy.  He  endeavors  so  far  to  identify  himself  with  the  contending 
parties  as  to  gain  a  clear  comprehension  of  their  motives,  but,  at  the  same  time,  h<r 
submits  their  actions  to  the  tests  of  a  cool  and  impartial  examination.*' — N.  Y.  Tribune. 


D.  APPLETON  &  CO.,  Publishers,  549  &  551  Broadway,  N.  Y, 


opinions  of  the  Press  on  the    International  Scientific  Series,^^ 

XIII. 

THE  DOCTRINE  OF 

Descent,  and  Darwinism. 

By  OSCAR  SCHMIDT, 
Professor  in  the  University  of  Strasburg. 

With  26  Woodcuts. 
I  vol.,  i2mo.    Cloth  Price,  $1.50. 

**  The  entire  subject  is  discussed  with  a  freshness,  as  well  as  an  elaboration  of  de- 
tail, that  renders  his  work  interesting  in  a  more  than  usual  degree.  The  facts  upon 
which  the  Darwinian  theory  is  based  are  presented  in  an  effective  manner,  conclusions 
are  ably  defended,  arid  the  question  is  treated  in  more  compact  and  available  style 
than  in  any  other  work  on  the  same  topic  that  has  yet  appeared.  It  is  a  valuable  ad- 
dition to  the  *  International  Scientific  Series,'  " — Boston  Post. 

*'  The  present  volume  is  the  thirteenth  of  the  *  International  Scientific  Series,'  and 
is  one  of  the  most  interesting  of  all  of  them.  The  subject-matter  is  handled  with  a 
great  deal  of  skill  and  earnestness,  and  the  courage  of  the  author  in  avowing  his  opin- 
ions is  much  to  his  credit.  .  .  .  This  volume  certainly  merits  a  careful  perusal." — 
Hartford  Evening  Post, 

**  The  volume  which  Prof  Schmidt  has  devoted  to  this  theme  is  a  valuable  contri- 
bution to  the  Darwinian  literature.  Philosophical  in  method,  and  eminently  candid, 
it  shows  not  only  the  ground  which  Darwin  had  in  his  researches  made,  and  conclu- 
sions reached  before  him  to  plant  his  theory  upon,  but  shows,  also,  what  that  theory 
really  is,  a  point  upon  which  many  good  people  who  talk  very  earnestly  about  the 
matter  are  very  imperfectly  informed." — Detroit  Free  Press. 

XIV. 

The  Chemistry  of  Light  and 
Photography ; 

In  its  Application  to  Art,  Science,  and  Industry. 

By  Dr.  HERMANN  VOGEI, 
Professor  in  the  Royal  Industrial  Academy  of  Berlin. 
With  100  Illustrations. 
i2mo  Price,  $2.00. 

**Out  of  Photography  has  sprung  a  new  science — the  Chemistry  of  Light — and,  in 
giving  a  popular  view  to  the  one.  Dr.  Vogel  has  presented  an  analysis  of  the  principles 
and  processes  of  the  other.  His  treatise  is  as  entertaining  as  it  is  instructive,  pleas- 
antly combining  a  history  of  the  progress  and  practice  of  photography — from  the  first 
rough  experiments  of  Wedgwood  and  Davy  with  sensitized  paper,  in  1802,  down  to 
the  latest  improvements  of  the  art— with  technical  illustrations  of  the  scientific  theories 
on  which  the  art  is  based.  It  is  the  first  attempt  in  any  manual  of  photography  to  set 
forth  adequately  the  just  claims  of  the  invention,  both  from  an  artistic  and  a  scientific 
point  of  view,  and  it  must  be  conceded  that  the  effort  has  been  ably  conducted."-^ 
Chicago  Tribune. 

D.  APPLETON  &  CO.,  Publishers,  549  &  551  Broadway,  N.  Y. 


opinions  of  the  Press  on  the  ^''International  Scientific  Series,"*^ 


XV. 

Fungi ; 

THEIR  NATURE,  INFLUENCE,  AND  USES. 
By  M.  C.  COOKE,  M.A,,  LL.  D. 
Edited  by  Rev.  M.  J.  BERKELEY,  M.  A.,  F.  L.  S. 
With  109  Illustrations.    Price,  $1.50. 

"Even  if  the  name  of  the  author  of  this  work  were  not  deservedly  eminent,  that  of 
the  editor,  who  has  long  stood  at  the  head  of  the  British  fungologists,  would  be  a  suf- 
ficient voucher  for  the  accuracy  of  one  of  the  best  botanical  monographs  ever  issued 
from  the  press.  .  .  .  The  structure,  germination,  and  growth  of  all  these  widely-dif- 
fused organisms,  their  habitats  and  influences  i^"  good  and  evil,  are  systematically 
described." — New  York  World. 

**  Dr.  Cooke's  book  contains  an  admirable  risuwd of  what  is  known  on  the  struct- 
ure, growth,  and  reproduction  of  fungi,  together  with  ample  bibliographical  references 
to  original  sources  of  information." — London  AthericBum. 

"The  production  of  a  work  like  the  one  now  under  review  represents  a  large 
amount  of  laborious,  difficult,  and  critical  work,  and  one  in  which  a  serious  slip  or  fatal 
error  would  be  one  of  the  easiest  matters  possible,  but,  as  far  as  we  are  able  to  judge, 
the  new  hand-book  seems  in  every  way  well  suited  to  the  requirements  of  all  beginners 
in  the  difficult  and  involved  study  of  fungology." — The  Gardener' s  Chro7iicle  {Lon- 
don). 


XVI. 

The  Life  and  Growth  of  Language: 

AN    OUTLINE    OF    LINGUISTIC  SCIENCE. 
By  WILLIAM  D WIGHT  WHITNEY, 

Professor  of  Sanskrit  and  Comparative  Philology  in  Yale  College. 
I  vol.,  i2mo.    Cloth.    Price,  $1.50. 

**  Prof  Whitney  is  to  be  commended  for  giving  to  the  public  the  results  of  his  ripe 
scholarship  and  unusually  profound  researches  in  simple  language.  He  draws  illus- 
trations and  examples  of  the  principles  which  he  wishes  to  impact,  from  common  Hfe 
and  the  words  in  frequent  use. 

The  topics  discussed  in  this  volume  are,  for  the  most  part,  those  which  have 
been  already  treated  by  other  writers  on  philology,  and  even  by  the  author  himself,  in 
his  volume  on  'Language,  and  the  Study  of  Language,'  published  a  few  years  ago, 
and,  though  many  of  the  truths  here  set  forth  are  those  with  which  students  in  the 
same  line  of  investigation  are  generally  familiar,  all  will  rejoice  to  see  them  restated  in 
such  a  fresh  and  simple  way. 

*'This  work,  while  valuable  to  scholars,  will  be  interesting  to  every  one." — The 
Churckjnan. 

"  This  work  is  an  important  contribution  to  a  science  which  has  advanced  steadily 
under  conditions  that  appear  constantly  to  throw  an  increasing  light  on  difficult  ques- 
tions, and  at  each  step  clear  the  way  for  further  discoveries." — Chicago  hiter-Occan. 

Prof.  Whitney  is  undoubtedly  one  of  the  foremost  of  English-speaking  philologists, 
and  occupies  an  enviable  position  in  the  wider  circle  of  European  students  of  language. 

**  His  style,  clear,  simple,  picturesque,  abounding  in  striking  illustrations,  and  apt 
in  comparisons,  is  admiral)ly  fitted  to  he  the  vehicle  of  a  popular  treatise  like  the  work 
under  consideration." — Poi  iland  Daily  Press. 


D.  APPLETON  &  CO.,  Publishers,  549  &  551  Broadway,  N.  Y. 


1( 


